Topical Compositions Comprising Fipronil and Permethrin and Methods of Use

ABSTRACT

The subject matter disclosed herein is directed to stable, highly-effective topical formulations comprising permethrin, fipronil and a solvent system that is sufficient to solubilize these two active ingredients and limit degradation of fipronil to its sulfone, and their uses in topical applications on animals and the environment. Useful formulations comprise from about 30% to about 55% (w/w) permethrin and about 2 to 15% (w/w) fipronil and a solvent system that comprises N-methyl pyrrolidone and a glycol, glycol ether, glycol ester, fatty acid ester or neutral oil, wherein the N-methyl pyrrolidone and glycol, glycol ether, glycol ester, fatty acid ester or neutral oil are present in a weight:weight ratio of from about 1:2.0 to about 1:3.5, glycol, glycol ether, glycol ester, fatty acid ester or neutral oil to n-methyl pyrrolidone. These two actives when combined in the described amounts have been found to have unexpected enhanced repellent activity against stable fly. However, it is the formulations described herein that provide solvency and stability that maintain synergistic concentrations after application on an animal.

CROSS REFERENCE TO RELATED APPLICATIONS

This application claims the benefit of priority to U.S. ProvisionalPatent Application No. 61/602,472, filed 23 Feb. 2012, which isincorporated herein by reference in its entirety.

FIELD OF THE INVENTION

The subject matter disclosed herein is directed to pharmaceutical andveterinary formulations that provide an unexpected enhanced repellantactivity and superior parasiticidal efficacy. The solvent system of theformulations provide excellent solvency and reduce the amount offipronil sulfone formation over a wide range of conditions.

BACKGROUND OF THE INVENTION

Animals, such as mammals and birds are often susceptible to parasiteinfestations. These parasites may be ectoparasites, such as fleas,ticks, mosquitos, mites, flies, sand flies and lice and endoparasitessuch as roundworms, hookworms, flukes and heartworms. Domesticatedanimals including farm animals are particularly susceptible to parasiteinfections and infestations.

Parasitic diseases may be caused by either endoparasites orectoparasites. Endoparasites refer to those parasites living inside thebody of the host (such as in the stomach, lungs, heart, intestines,etc.). Ectoparasites are those parasites that live on the outer surfaceof the host but still draw nutrients from the host. Endoparasiticinfections may further be subdivided based on class of parasite involvedin the infection. These parasitic infections and infestations are oftenassociated with illness and death or reduced productivity.

Examples of endoparasites which infect animals include but are notlimited to gastro-intestinal parasites of the genera Ancylostoma,Necator, Ascaris, Strongyloides, Trichinella, Capillaria, Toxocara,Toxascaris, Trichuris, Enterobius, Haemonchus, Trichostrongylus,Ostertagia, Cooperia, Oesophagostomum, Bunostomum, Strongylus,Cyathostomum and Parascaris among others, and those that are found inthe blood vessels or other tissues and organs include Wuchereria,Brugia, Onchocerca, Dirofilaria and the extra-intestinal stages ofStrongyloides, Toxocara and Trichinella.

Ectoparasites which infest man and domestic animals include arthropods,such as ticks, fleas, mites, mosquitoes, lice, and the like andinfestations by these parasites can result in transmission of pathogensthat can cause serious and even fatal diseases.

Infestations by ectoparasitic arthropods including but not limited toticks, mites, lice, stable flies, horn flies, blowflies, face flies,fleas, mosquitoes and the like are also a serious problem. Infestationby these parasites may result not only in loss of blood and skinlesions, but also can interfere with normal eating habits thus causingweight loss. Ectoparasitic infestations of a host can also result intransmission of pathogenic agents that cause serious diseases includingbut not limited to encephalitis, anaplasmosis, Babesiosis, RockyMountain spotted fever, Lyme disease, ehrlichiosis, West Nile virus,swine pox, malaria, yellow fever, and others, many of which can be fatalto the host. Animals may be infected by several species of parasites atthe same time since infection by one parasite may weaken the animal andmake it more susceptible to infection by a second species of parasites.

A parasite which is prevalent among domesticated animals is Stomoxyscalcitrans (stable fly). Stable flies will feed on blood frompractically any warm blooded animal, including humans, pets, andlivestock. During periods of high stable fly activity, humans can beseverely annoyed and this insect has been called the biting house fly.Individual flies may feed more than once per day. Peaks of feedingactivity commonly occur during the early morning and again in the lateafternoon. Stable flies prefer feeding on lower parts of the hosts suchas the legs.

Both male and female stable flies feed on blood. The female requiresblood meals to produce viable eggs. Females deposit their eggs in avariety of decaying animal and plant wastes. Stable flies breed in soggyhay, grasses or feed, piles of moist fermenting weed or grass cuttings,spilled green chop, peanut litter, and seaweed deposits among beaches,in soiled straw bedding and sometimes in hay ring feeding sites. Eachfemale fly may lay 500-600 eggs in 4 separate batches. Eggs hatch in 2-5days into larvae which feed and mature in 14-26 days. The average lifecycle is 28 days, ranging from 22-58 days, depending on the weatherconditions.

Farm animals can be severely affected by the stable fly. Since the flytakes blood meals, animals are weakened from blood loss and continualirritation. Animals such as swine, cattle, and horses infested withstable flies may show reduced weight gains, among other harmful effects.While one stable fly does not cause significant damage, 50-100 of theseblood-sucking pests together with 500 horn flies cause a significantloss of blood. This can result in a significant loss of milk and beefproduction from animals each year.

Another important parasite is the tick genus Rhipicephalus, especiallythose of the species microplus (cattle tick), decoloratus and annulatus.Ticks, such as Rhipicephalus microplus, are particularly difficult tocontrol because they live in the pasture where the farm animals graze.Other important parasites of cattle and sheep are listed as follows:myiases such as Dermatobia hominis (known as Berne in Brazil) andCochliomyia hominivorax (greenbottle); sheep myiases such as Luciliasericata, Lucilia cuprina (known as blowfly strike in Australia, NewZealand and South Africa). These are flies whose larva constitute theanimal parasite; flies proper, namely those whose adult constitutes theparasite, such as Haematobia irritans (horn fly); lice such asLinognathus vitulorum, etc.; and mites such as Sarcoptes scabiei andPsoroptes ovis.

More prevalent among domesticated animals, such as cats and dogs, arethe following ectoparasites: cat and dog fleas (Ctenocephalides felis,C. canis. and the like), ticks (Rhipicephalus spp., Ixodes spp.,Dermacentor spp., Amblyoma spp. and the like), and mites (Demodex spp.,Sarcoptes spp., Cheyletiella spp., Otodectes spp. and the like), lice(Trichodectes spp., Lignonathus spp., and the like), mosquitoes (Aedesspp., Culex spp., Anopheles spp., and the like) and flies (Hematobiaspp., Musca spp., Stomoxys spp., Dermatobia sp., Cochliomyia spp., andthe like).

Fleas are a particular problem because not only do they adversely affectthe health of the animal or human, but they also cause a great deal ofpsychological stress. Moreover, fleas are also vectors of pathogenicagents in animals, such as dog tapeworm (Dipylidium caninum), andhumans.

Similarly, ticks are also harmful to the physical and psychologicalhealth of the animal. However, the most serious problem associated withticks is that they are the vector of pathogenic agents, agents whichcause diseases in both humans and animals. Major diseases which arecaused by ticks include borrelioses (Lyme disease caused by Borreliaburgdorferi), Babesioses (or piroplasmoses caused by Babesia sp.) andrickettsioses (also known as Rocky Mountain spotted fever). Ticks alsorelease toxins which cause inflammation or paralysis in the host.Occasionally, these toxins are fatal to the host.

Moreover, mites and lice are particularly difficult to combat sincethere are very few active substances which act on these parasites andthey require frequent treatment.

The above list is not exhaustive and other ectoparasites are well knownin the art to be harmful to animals and humans.

While it is known in the art that it is sometimes possible to combinevarious parasiticides in order to broaden the parasiticidal spectrum, itis not possible to predict, a priori, with any reasonable certaintywhich combinations will work for a particular animal or disease state.For this reason, the results of various combinations are not alwayssuccessful and there is a need in the art for more effectiveformulations which may be easily administered to the animal and have therequired solvency, stability and bioavailability.

Formulations comprising different actives are especially difficult tosuccessfully formulate because of the challenges in achieving therequired solvency, stability and bioavailability. Thus, there is a needin the art for combination antiparasitic formulations that meet therequired solvency, stability and bioavailability of the parasiticides tobe formulated therein.

SUMMARY OF THE INVENTION

The subject matter disclosed herein is directed to stable,highly-effective topical formulations comprising a pyrethroid, such aspermethrin, a N-arylpyrazole, such as fipronil and a solvent system thatis sufficient to solubilize these two active ingredients and limit theoxidation of fipronil to fipronil sulfone, but that is safe for topicalapplication. In one embodiment, the topical formulation is in the formof a spot-on formulation. In another embodiment, the topical formulationis in the form of a pour-on formulation. In one embodiment, formulationscomprise about 30 to about 55% (w/w) permethrin and about 2 to 15% (w/w)fipronil and a solvent system that comprises N-methyl pyrrolidone (NMP)and a glycol, glycol ether, glycol ester, fatty acid ester or neutraloil. In some embodiments, the formulations of the invention comprise atleast about 40% (w/w) permethrin and at least about 5% (w/w) fipronil ina solvent system comprising NMP and a glycol, glycol ether, glycolester, fatty acid ester or neutral oil, wherein the NMP and glycol,glycol ether, glycol ester, fatty acid ester or neutral oil are presentin a ratio of from about 1:2.0 to about 1:3.5 (weight:weight), glycol,glycol ether, glycol ester, fatty acid ester or neutral oil to NMP. Inother embodiments, the ratio of glycol, glycol ether, glycol ester,fatty acid ester or neutral oil to NMP is about 1:2.0 to about 1:3.0,about 1:2.5 to about 1:3.5, or about 1:2.5 to about 1:3.0(weight:weight).

These two actives when combined in the described amounts in the solventsdescribed herein have been found to have excellent parasiticidalactivity and unexpected enhanced repellency against stable flies.However, it is the particular combination of solvents in theformulations described herein that provide the required solvency andphysical stability and limit the oxidation of fipronil to its sulfonethat maintain concentrations of the actives that result in theunexpected enhanced repellency and superior efficacy against stableflies after application on an animal. Additionally, as described herein,the selection of the diluent to be combined with NMP is critical tolimiting the oxidation of fipronil to fipronil sulfone in theformulations and maintaining effective concentrations.

In some embodiments of the invention, additional active agents may becombined in the topical formulations comprising fipronil and permethrin.The additional active agents include, but are not limited to, anavermectin, a milbemycin, a spinosyn, a spinosoid, a benzimidazole,levamisole, pyrantel, morantel, praziquantel, closantel, clorsulon, anamino acetonitrile active agent, an insect growth regulator, aneonicotinoids or an aryloazol-2-yl cyanoethylamino active agent, or acombination of thereof.

The subject matter disclosed herein is also directed to methods of useof the formulations to combat infestations of ectoparasites, including,but not limited to, fleas, ticks, mosquitoes, and various types ofparasitic flies, such as stable flies, on or around mammals or birds andin the environments where they reside. The methods compriseadministering an effective amount of a formulation described herein, inparticular a spot-on or pour-on formulation, to the mammal or bird.Animals include mammals, such as dogs, zebras and horses, cattle, pigs,yaks, rodents, deer, goats, sheep and llamas and birds, such aschickens, turkeys and quail. Environments include animal houses, such asdog bedding, horse stables and chicken litter.

The present subject matter is also directed to a method of preparing thecompositions described herein. The method comprises contacting one orboth of the actives with one or more of the solvents selected fromN-methyl pyrrolidone and a glycol, glycol ether, glycol ester, fattyacid ester or neutral oil, wherein the final formulation containsN-methyl pyrrolidone and glycol, glycol ether, glycol ester, fatty acidester or neutral oil. In one embodiment, the glycol, glycol ether,glycol ester, fatty acid ester or neutral oil and NMP are combined in aratio of from about 1:2.0 to about 1:3.5, about 1:2.0 to about 1:3.0,about 1:2.5 to about 1:3.5, or about 1:2.5 to about 1:3.0(weight:weight), glycol, glycol ether, glycol ester, fatty acid ester orneutral oil to NMP. The formulations described herein are stable andexhibit unexpected enhanced repellency and efficacy against stableflies.

BRIEF DESCRIPTION OF THE DRAWINGS

The following detailed description, given by way of example, but notintended to limit the invention solely to the specific embodimentsdescribed, may best be understood in conjunction with the accompanyingdrawings, in which:

FIG. 1 depicts sulfone formation at 50° C. for formulations containingN-methylpyrrolidone (NMP) and diethyleneglycol monoethyl ether (DGME,TRANSCUTOL®) or MIGLYOL® 812 (a neutral oil). The data show that theDGME-containing formulation contains more than 10% higher concentrationof fipronil sulfone compared to the neutral oil-containing formulation.Each formulation contains about 6% (w/w) fipronil, 45% (w/w) permethrinand 33% (w/w) NMP. Formulation A contains (DGME), QS to 100% (˜14-15%w/w); formulation B contains MIGLYOL® 812, QS to 100% (˜14-15% w/w). Theformulations in this study do not contain an added antioxidant.

FIG. 2 depicts formulations which all contain MIGLYOL® 812 and either noantioxidant or a different level of antioxidant. The data show that allformulations have fipronil sulfone levels below about 2.950%. Theformulation containing 0.10% BHT has a fipronil sulfone level of about2.800%.

FIG. 3 depicts sand fly repellency of formulations containingfipronil/permethrin and NMP/DGME or NMP/MIGLYOL® 812.

FIG. 4 depicts efficacy against fleas of formulations containingfipronil/permethrin and NMP/DGME, NMP/MIGLYOL® 812 orN,N-dimethyldecanamide (DMDA)/butyl CELLOSOLVE™ acetate (BCA), orpermethrin alone. NMP/DGME formulations are more susceptible to fipronildegradation

FIG. 5 depicts efficacy against ticks of formulations containingfipronil/permethrin and NMP/DGME, NMP/MIGLYOL® 812 or DMDA/BCA, orpermethrin alone.

FIG. 6 depicts efficacy against stable fly on rats with a formulationcomprising permethrin alone or fipronil/permethrin.

FIG. 7 depicts repellency against stable fly on rats with a formulationcomprising permethrin alone or fipronil/permethrin.

FIG. 8 depicts repellency against stable fly on dogs of a formulationcomprising fipronil and permethrin one hour post exposure.

FIG. 9 depicts the efficacy against stable fly on dogs of a formulationcomprising fipronil and permethrin one hour post exposure.

FIG. 10 depicts the repellency against Culex pipiens mosquitoes of aformulation comprising fipronil and permethrin one hour post exposure.

DETAILED DESCRIPTION

The presently disclosed subject matter will be described more fullyherein. However, many modifications and other embodiments of thepresently disclosed subject matter set forth herein will come to mind toone skilled in the art to which the presently disclosed subject matterpertains having the benefit of the teachings presented in the foregoingdescription. Therefore, it is to be understood that the presentlydisclosed subject matter is not to be limited to the specificembodiments disclosed and that modifications and other embodiments areintended to be included within the scope of the appended claims.

It is well-known that formulating an active into a spot-on or pour-onformulation is challenging. Even more challenging is successfullyformulating two different actives into a composition, whereby bothactives remain in solution and are available in the desired amounts.Since it has been shown herein that the combination has an unexpectedenhanced repellency against stable flies, it is critical that theformulation provide the required solubility and physical and chemicalstability to ensure that the concentrations resulting in enhancedrepellency are maintained even after administration of the formulationto the animal.

Disclosed herein is a unique combination of a solvent system for usewith the combination of permethrin and fipronil, whereby the formulationcomponents are present in specified ranges and particular ratiosrelative to one another. Such enumerated ratios are not broadlygeneralized amounts, but are specific values of effective ranges andratios for fipronil and permethrin. Advantageously, the solvent systemsdescribed herein solubilize a high concentration of permethrin and arecompatible with fipronil. Consequently, the actives stay in solution,are not substantially degraded and are available to combat pestinfestations at effective concentrations at the loci where infestationscan occur. As used herein, the term “physical stability” and “physicallystable” refers to a property of a formulation wherein no significantamount permethrin or fipronil crystals form at low temperatures(including −20° C., 4° C. and 10° C.), the active ingredients remain insolution even after application. The physical stability of thecompositions may be described in terms of crystal formation when theformulation is stored at a particular temperature for a period of days,weeks, months or even years. The solvent system also provides therequired safety profile for its intended uses.

Formulations that contain permethrin and an additional active ingredientoften suffer from physical instability. Permethrin, especially in highconcentrations above 20% (w/w), tends to form crystals in theformulation. When a second active is added to the formulation, thephysical instability problems can increase exponentially since thephysical properties and requirements for formulation of an additionalingredient must also be considered. The second preferred activedisclosed herein is fipronil. However, when fipronil is combined withN-methylpyrrolidone (NMP), which is an effective solvent for highconcentrations of permethrin, significant levels of oxidation offipronil to fipronil sulfone is observed.

Fipronil sulfone is also a principal in vivo metabolite of fipronil. Ithas the following structure:

Fipronil used to prepare the formulations can contain fipronil sulfoneat the time of formulation. In other words, it is possible that usefulgrades of fipronil have some amount of fipronil sulfone already presentas a byproduct of its preparation. In one embodiment fipronil has belowabout 3.5% fipronil sulfone at the time of formulation with NMP. Somegrades of fipronil may have lower levels of fipronil sulfone, even 0%.However, once fipronil is contacted with NMP, it is believed thatfipronil sulfone may continue to form in solution. Further, it issignificantly more costly to use fipronil that is completely free offipronil sulfone. However, the formulation described hereinadvantageously minimize or substantially slow further fipronil sulfoneformation compared to formulations with other common solvents used intopical compositions no matter how much is present at the time offormulation.

The solvent system described herein was designed after the finding thatfipronil is susceptible to fipronil sulfone formation in the presence ofNMP. While not being bound to any particular theory, it is believed thatperoxides present in NMP can facilitate the formation of fipronilsulfone. Furthermore, it is known that NMP can generate peroxides in thepresence of oxygen (see Drago and Riley, J. Am. Chem. Soc. 1990, 112,215-218; Reynolds et al., Journal of Pharmaceutical Sciences, 2012,101(2), pp. 761-776). Thus, it has been found that NMP, while a usefulsolvent, poses technical hurdles when used as a solvent for fipronil ina concentrated topical formulation, such as a spot-on formulation. Theoxidation of fipronil in NMP must be addressed since NMP is an effectivesolvent for permethrin at the concentration levels present in theformulations disclosed herein. However, the required amount of NMP inthe formulations of the invention was unexpectedly observed to result inthe oxidation of fipronil to fipronil sulfone at levels that were higherthan desired.

N-methylpyrrolidone (NMP) is a chemical compound with a 5-memberedlactam structure. It is a clear to slightly yellow liquid soluble inwater and conventional organic solvents. It is a polar aprotic solventand a very weak base. Other names for this compound are:1-methyl-2-pyrrolidone, N-methyl-2-pyrrolidinone, and m-pyrrole, andPHARMASOLVE®. Despite its usefulness, its use for topical applicationmay pose problems for the skilled formulator. In some cases NMP maycause skin irritation, redness or dermatitis. Accordingly, although itis acceptable in topical formulations, it is preferred that the level beas low as possible.

The oxidation of fipronil to fipronil sulfone was observed at higherthan desired levels when NMP was diluted with traditional solvents usedin combination with fipronil in topical formulations, such asdiethyleneglycol monoethyl ether (TRANSCUTOL® or DGME). Diethyleneglycolmonoethyl ether is commonly used in many topical formulations and hasbeen found to be a very effective solvent for use in formulating topicalfipronil formulations. When DGME and other diluents are used incombination with NMP, the amount of NMP can be reduced so long as thenecessary physical properties of the composition remain unaltered.However, it has been found that when NMP is combined with co-solventsnormally used with fipronil, such as DGME, fipronil is oxidized to itssulfone at higher than desired rates. Consequently, to minimize theformation of fipronil sulfone, the use of an antioxidant at about10-times the level of antioxidant that is effective in the formulationsdescribed herein was required.

It has been surprisingly found that a solvent system containing NMP incombination with a neutral oil, including MIGLYOL® 812 (caprylic/caprictriglyceride), results in significantly lower sulfone formation comparedwith other solvents typically used in fipronil topical formulations.Neutral oils are known in the art. They are light to colorless liquidsof neutral odor and taste.

MIGLYOL®-type neutral oils are clear, slightly yellowish esters ofsaturated coconut and palm kernel oil-derived caprylic and capric fattyacids and glycerin or propylene glycol. Some neutral oils aretriglycerides of fractionated plant fatty acids with chain lengths of C₈to C₁₀. Two commercially available products are known as MIGLYOL® 810and MIGLYOL®812. Another useful neutral oil is a triglyceride offractionated plant fatty acids with chain lengths of C₈ and C₁₀ combinedwith linoleic acid (about 4-5%). A commercially available product isknown as MIGLYOL® 818. Another useful neutral oil is a glycerin ester offractionated plant fatty acids with chain lengths of C₈ and C₁₀ combinedwith succinic acid. A commercially available product is known asMIGLYOL® 829. Another useful neutral oil is a propylene glycol diesterof saturated plant fatty acids with chain lengths of C₈ and C₁₀ combinedwith succinic acid. A commercially available product is known asMIGLYOL® 840 (propylene glycol dicaprylate/dicaprate). Other fatty acidsinclude stearyl stearate, palmitate, and myristate. Other examples ofsuitable neutral oils and fatty acid esters include hydrocarbonaceousvegetable oils, such as liquid triglycerides of fatty acids comprisingfrom 4 to 24 carbon atoms (such as triglycerides, heptanoic acid andoctanoic acid), sunflower oil, maize oil, soybean oil, gourd oil, grapeseed oil, sesame oil, hazelnut oil, apricot oil, macadamia oil, castoroil, avocado oil, jojoba oil and karite butter; synthetic esters, suchas synthetic esters of fatty acids such as, for example, purcellin oil,isononyl isononanoate, isopropyl myristate, 2-ethylhexyl palmitate,2-octyldodecyl stearate, 2-octyldodecyl erucate, and isostearylisostearate. As described fully herein, the requirement of the diluentis that it does not substantially introduce additional peroxide speciesinto the NMP/co-solvent system. Consequently, fipronil sulfone formationis mitigated and substantially lower levels of antioxidants can be usedto yield long-term stability of the permethrin, fipronil combinationformulation.

The fipronil/permethrin compositions described herein containing NMP incombination with neutral oils will have significantly less fipronilsulfone growth compared to fipronil/permethrin compositions that use adifferent NMP/co-solvent system. It is useful that the presentcompositions inhibit fipronil sulfone formation such that the level offipronil sulfone present at about 3 months after formulation has notincreased by more than about 50% of the original fipronil sulfonepresent at the time of formulation, e.g., the amount present in neatfipronil raw material, as measured by area % of the fipronil sulfonepeak relative to the fipronil peak by HPLC. In one embodiment, thesolvent system of the invention inhibit the formation of fipronilsulfone to an extent that at three months after formulation, thefipronil sulfone has not increased by more than about 45% of theoriginal fipronil sulfone present at the time of formulation; or thefipronil sulfone has not increased by more than about 40% of theoriginal fipronil sulfone present at the time of formulation; or thefipronil sulfone has not increased by more than about 35% of theoriginal fipronil sulfone present at the time of formulation; or thefipronil sulfone has not increased by more than about 30% of theoriginal fipronil sulfone present at the time of formulation; or thefipronil sulfone has not increased by more than about 25% of theoriginal fipronil sulfone present at the time of formulation. In anotherembodiment, at three month after formulation, the fipronil sulfone hasnot increased by more than about 20% of the original fipronil sulfonepresent at the time of formulation; or the fipronil sulfone has notincreased by more than about 15% of the original fipronil sulfonepresent at the time of formulation. In yet another embodiment, at threemonth after formulation, the fipronil sulfone has not increased by morethan about 10% of the original fipronil sulfone present at the time offormulation.

The purpose of the antioxidant in the compositions is largely to combatany peroxides found in NMP. It has been found that when a diluent iscarefully selected to limit additional introduction of harmful oxidativespecies, such as peroxide species, into the composition, the amount ofantioxidant can be preferably kept at GRAS (generally regarded as safe)levels or substantially lower. In some embodiments, an effective amountof antioxidant is not more than about 0.25% (w/w). In anotherembodiment, the antioxidant is present at a concentration of not morethan about: 0.2% (w/w); 0.19% (w/w); 0.18% (w/w); 0.17% (w/w); 0.16%(w/w); 0.15% (w/w); 0.14% (w/w); 0.13% (w/w); 0.12% (w/w) or 0.11%(w/w). In still another embodiment, the antioxidant is present at aconcentration of not more than about 0.1% (w/w).

Another problem with concentrated topical formulations is that anyamount of crystals in the formulation may lead to a concentration ofactive(s) in the precipitated material that is too high for safe,point-of-application topical use. Further, since precipitationeffectively removes active(s) from solution, the concentration ofactive(s) remaining in solution can be too low, and thus not provide theefficacy described herein. Additionally, once crystals begin to form,they behave as seeds for further crystal formation. This process resultsin increasing precipitation of components out of the solvent system, anddeleterious effects on the concentration of active(s) in theformulation.

It is evident that should one or both of the actives form crystals orotherwise precipitate from the solution either before or afterapplication, the relative amounts of the active agents would be altered.Accordingly, the amounts of actives would be expected to deviate fromthe amounts required for enhanced repellent activity. Therefore, theproperty of the present compositions whereby each active remains insolution even after application on the animal also contributes to theenhanced activity against stable flies observed. As reported inWO2007/143298, incorporated herein in its entirety, it is difficult toform a high concentration solution of permethrin. However, the presentcompositions contain about 30% (w/w) permethrin at the lowestconcentration, and in some embodiments contain about 40 to about 45(w/w). The difficulty of successfully formulating permethrin in theseconcentrations is exponentially greater when formulating a second,different active. This is especially true if the actives are ofdifferent physical properties. Surprisingly, the compositions describedherein have been shown to exhibit surprising enhanced repellency andexcellent insecticidal efficacy against stable flies when applied to alocalized area on an animal. Thus, the enhanced repellency observedwould be expected to result from the concentration of each active as thecomposition spreads over the animal's coat and skin. The unexpectedenhanced repellency would not be limited to the loci where thecomposition is directly applied to the animal, and this is important foreffective control of stable flies and other ectoparasites since theseectoparasites will feed on different locations on the animal. Theformulations described herein provide effective amounts of the activeagents of the compositions even as the composition translocates over theanimal.

Although many solvent systems are available, a skilled formulator cannotpredict which system will effectively work for a given combination ofactive ingredients. However, the presently described solvent systemsolves the problems of formulating fipronil and permethrin together ineffective amounts that result in surprising enhanced repellency againstcertain parasitic flies, including stable flies, wherein the effectiveconcentrations of fipronil and permethrin resulting in the enhancedrepellency are maintained.

Fipronil(5-amino-3-cyano-1-(2,6-dichloro-4-trifluoromethylphenyl)-4-trifluoromethylsulfinylpyrazole)is a 1-N-arylpyrazole insecticide. It has the following structure:

It is a member of a class of chemicals that are well known in the art,as well as methods for their use in controlling parasites includinginsect pests, such as fleas, stable flies, horn flies, or mosquitoes,and acarid pests, such as ticks, mites, and lice, in mammals, such asdomesticated livestock or companion animals or birds, either alone or incombination with other pesticides such as insect growth regulators. See,e.g., EP-A-295,217, EP 295 177, EP-A-840-686, EP-A-352,944, WO 00/35844,WO 98/39972, U.S. Pat. Nos. 5,122,530 5,236,938, 5,232,940, 5,576,4295,814,652, 5,567,429, 6,090,751 and 6,096,329. These references arehereby incorporated herein in their entirety. The compounds of thefamilies defined in these patents are extremely active. Fipronil isparticularly effective, but not exclusively effective, against fleas andticks. However, fipronil is not known to have any repellent activityagainst ectoparasites.

Permethrin is a member of the pyrethroids. It has the followingstructure:

The pyrethroids are a class of synthetically derived insecticides. Thesecompounds are particularly effective against the mosquito (Culex spp.)that transmits West Nile virus. Pyrethroids are structurally related tonaturally occurring pyrethrins, pyrethrin I and pyrethrin II. Syntheticpyrethroids include permethrin (U.S. Pat. No. 4,113,968), resmethrin,and sumithrin (U.S. Pat. Nos. 3,934,023 and 2,348,930). Permethrin iswell known for its repellent efficacy against various arthropods. Thesereferences are hereby incorporated herein in their entirety.

Combinations of fipronil and pyrethroids, such as permethrin, are known(in particular, EPA Registration submission materials available from aPDMS, searchable database; submitter name: Virbac AH, Inc.) The productsold by the trade name EFFITIX® was approved in the U.S. on Nov. 18,2011. A material safety data sheet (MSDS) of the product indicates thatit contains fipronil, permethrin and diethyleneglycol monoethyl ether.WO 2001/065941 A1 and EP 1013170 A1 propose the combination of anN-arylpyrazole and a pyrethroid in applications against plant pests. JP11049618 A2 uses similar mixtures to prevent feeding damage on timberconstructions. WO 95/22902 A1 uses such mixtures for the direct controlof termites. GB2396557 A1 teaches that the treatment of ectoparasiteswith mixtures of N-arylpyrazoles and pyrethroids (if appropriate alsowith addition of synergists, such as MGK264 or piperonyl butoxide) ispossible when concentrated powder formulations are used. All of thesereferences are hereby incorporated herein in their entirety.

However, in order for the actives to function, it is critical to employa solvent system that provides the necessary solubility and chemical andphysical stability to ensure that effective concentrations responsiblefor the enhanced repellent efficacy remain in solution afteradministration and up until the actives cover the animal effectively orreach a locus of infestation. There has been no report of theformulation components required to successfully formulate fipronil andpermethrin in combination where permethrin is present at highconcentrations in a spot-on or pour-on formulation that maintainsconcentrations under a variety of conditions that result in enhancedrepellency against certain ectoparasites including, but not limited to,stable flies. Even skilled formulators cannot predict with reasonablecertainty which combinations and solvents will ultimately work together.

In an embodiment, the present subject matter is directed to aformulation comprising permethrin in a concentration of from about 30%w/w to about 55% w/w; fipronil in a concentration of from about 2% w/wto about 15% w/w; and a) one or more neutral oils and b) N-methylpyrrolidone (NMP).

In another embodiment, the invention provides a formulation comprisingpermethrin in a concentration of from about 30% w/w to about 55% w/w;fipronil in a concentration of from about 2% w/w to about 15% w/w; anda) one or more neutral oils and b) NMP, wherein the a) one or moreneutral oils and b) NMP are present in a weight:weight ratio of fromabout 1:18 to about 1:2.8, neutral oil(s) to NMP. In this embodiment, ifan antioxidant is present, it may be present in a concentration of notmore than about 0.25% (w/w).

In still another embodiment, the invention provides a formulationcomprising permethrin in a concentration of from about 30% w/w to about55% w/w; fipronil in a concentration of from about 2% w/w to about 15%w/w; and a) one or more neutral oils and b) NMP, wherein the a) one ormore neutral oils and b) NMP are present in a weight:weight ratio offrom about 1:2.0 to about 1:3.5, neutral oil(s) to NMP. In thisembodiment, if an antioxidant is present, it may be present in aconcentration of not more than about 0.25% (w/w).

In one embodiment, the invention provides a formulation comprisingpermethrin in a concentration of at least about 40% w/w; fipronil in aconcentration of at least about 5% w/w; and a) one or more neutral oilsand b) NMP, wherein the a) one or more neutral oils and b) NMP arepresent in a weight:weight ratio of from about 1:1.18 to about 1:2.8 orabout 1:2.0 to about 1:3.5, neutral oil(s) to NMP. In this embodiment,if an antioxidant is present, it may be present in a concentration ofnot more than about 0.25% (w/w).

In another embodiment, the invention provides a formulation comprisingpermethrin in a concentration of at least about 40% w/w; fipronil in aconcentration of at least about 5% w/w; and a) one or more neutral oilsand b) NMP, wherein the a) one or more neutral oils and b) NMP arepresent in a ratio of from about 1:2.5 to about 1:3.5, neutral oil(s) toNMP. In this embodiment, if an antioxidant is present, it may be presentin a concentration of not more than about 0.25% (w/w).

In still another embodiment, the invention provides a formulationcomprising permethrin in a concentration of from about 40% w/w to about55% w/w; fipronil in a concentration of from about 5% w/w to about 15%w/w; and a) one or more neutral oils and b) NMP, wherein the a) one ormore neutral oils and b) NMP are present in a weight:weight ratio offrom about 1:1.8 to about 1:2.8 or about 1:2.0 to about 1:3.5, neutraloil(s) to NMP. In this embodiment, if an antioxidant is present, it maybe present in a concentration of not more than about 0.25% (w/w).

In still another embodiment, the invention provides a formulationcomprising permethrin in a concentration of from about 40% w/w to about55% w/w; fipronil in a concentration of from about 5% w/w to about 15%w/w; and a) one or more neutral oils and b) NMP, wherein the a) one ormore neutral oils and b) NMP are present in a ratio of from about 1:2.5to about 1:3.5, neutral oil(s) to NMP. In this embodiment, if anantioxidant is present, it may be present in a concentration of not morethan about 0.25% (w/w).

In yet another embodiment, the invention provides a formulationcomprising permethrin in a concentration of from about 40% w/w to about50% w/w; fipronil in a concentration of from about 5% w/w to about 10%w/w; and a) one or more neutral oils and b) NMP, wherein the a) one ormore neutral oils and b) NMP are present in a ratio of from about 1:2.5to about 1:3.0, neutral oil(s) to NMP. In this embodiment, if anantioxidant is present, it may be present in a concentration of not morethan about 0.25% (w/w).

In one embodiment of the invention, the level of fipronil sulfone in thecompositions described herein is below about 6% by area relative to thearea of the fipronil peak measured by high pressure liquidchromatography (HPLC) at about three months after formulation. In otherembodiments fipronil sulfone levels will be below about 5% at aboutthree months after formulation or below about 4% at about three monthsafter formulation. In still other embodiments fipronil sulfone levelswill be below about 3.5% at about three months after the formulation isprepared. In another embodiment, the fipronil sulfone level will bebelow about 3.2% at about three months formulation, below about 3.1% atabout three months formulation, below about 3.0% at about three monthsformulation, below about 2.9% at about three months after formulation orbelow about 2.8% at about three months formulation. In otherembodiments, fipronil sulfone levels will be below about 2.7% at aboutthree months after formulation, below about 2.6% at about three monthsformulation, or below about 2.5% at about three months afterformulation. In yet other embodiments, fipronil sulfone levels will bebelow about 2.4% at about three months after formulation, below about2.3% at about three months after formulation, below about 2.2% at aboutthree months after formulation. In yet another embodiment, fipronilsulfone levels will be below about 2.1% at about three months afterformulation or below about 2.0% at about three months formulation. Asused herein, “about three months after formulation” means a period offrom 10 weeks to 14 weeks from the time that the fipronil is contactedwith other components in the formulation. Typically, it is the time fromcontact of the fipronil with NMP. The fipronil sulfone level isdetermined by HPLC relative to the area of the fipronil peak in thesample.

It is the ratio of one or more neutral oils to NMP that provides uniquestability of the two active ingredients, permethrin and fipronil in thepresent formulations, particularly since the neutral oil(s) will beselected so as to not introduce significant amounts of peroxide species.Unlike distinct formulations known in the art for different actives, theformulations disclosed herein are capable of solubilizing permethrin andinhibiting or preventing its crystallization, while also limiting theformation of fipronil sulfone in solution. Consequently, both activeingredients are available in the desired concentrations, which are shownherein to have unexpected enhanced efficacy against stable flies.

A useful concentration of fipronil in the compositions is from about 2%(w/w) to about 15% (w/w). In one embodiment, fipronil is present in aconcentration of from about 3% (w/w) to about 10% (w/w). In anotherembodiment, fipronil is present in a concentration of from about 4%(w/w) to about 8% (w/w). In yet another embodiment fipronil is presentat a concentration of about 6% (w/w).

A useful concentration of permethrin in the compositions of theinvention is from about 35% (w/w) to about 50% (w/w). In one embodiment,permethrin is present at a concentration of from about 40% (w/w) toabout 48% (w/w). In another embodiment, permethrin is present in aconcentration of about 42% (w/w) to about 47% (w/w). In still anotherembodiment, permethrin is present at a concentration of about 45% (w/w).

In some embodiments, the invention provides the following formulations:Formulation A) fipronil about 6% (w/w); permethrin about 45% (w/w); NMPabout 35% (w/w); DGME (diethyleneglycol monoethyl ether or TRANSCUTOL®)in an amount sufficient to complete the formulation (QS or quantumsufficit) about 10-15% (w/w) and about 0.1% (w/w) BHT; Formulation B)fipronil about 6% (w/w); permethrin about 45% (w/w); NMP about 35%(w/w); MIGLYOL® 812 in an amount sufficient to complete the formulation(QS) about 10-15% (w/w) and about 0.1% (w/w) BHT. In some embodiments,the amount of DGME in Formulation A) will be about 12-14%, about 12%(w/w), about 13% (w/w) or about 14% (w/w). In other embodiments, theamount of MIGLYOL® in Formulation B) will be about 12% (w/w), about 13%(w/w) or about 14% (w/w).

Dosages of the actives can be readily determined by those of skill inthe art. However, in general terms, a dose of from about 0.001 to about100 mg per kg of body weight, more typically from about 0.01 mg to about50 mg/kg of body weight, given as a single dose or in divided doses fora period of from 1 to 5 days may be satisfactory but, of course, therecan be instance where higher or lower dosage ranges are indicated andsuch are within the scope of this specific administration period for aparticular situation. It will be understood that the dosing volume of aformulation can also be determined and then adjusted as desired.

In some embodiments, the composition comprises fipronil at about 6.0%(w/w) and permethrin at about 45 (w/w). This composition also comprisesthe N-methyl pyrrolidone and neutral oil solvent system describedherein.

The neutral oil(s) and N-methyl pyrrolidone components can be quantifiedin terms of the relative amounts of each. In some embodiments, usefulweight:weight ratios of neutral oil(s) to NMP are from 1:1.8 to about1:3.5; or about 1:2.0 to about 1:3.5. In other embodiments, ratiosinclude from about 1:1.9 to about 1:2.7; about 1:2.0 to about 1:3.0; andabout 1:2.5 to about 1:3.0, neutral oil(s) to NMP. In other embodimentsratio values include: 1:1.9, 1:2, 1:2.1, 1:2.2, 1:2.4, 1:2.5, 1:2.6,1:2.7, 1:2.8, 1:2.9, 1:3.0, 1:3.1, 1:3.2, 1:3.3, 1:3.4 and 1:3.5,neutral oil(s) to NMP. In yet other embodiments, ratios include fromabout 1:2.2 to about 1:2.4, neutral oil(s) to NMP. In anotherembodiment, the ratio is about 1:2.3 to about 1:3.0, neutral oil(s) toNMP. These ratios are relative amounts, but are suitably based on auseful amount of neutral oil(s) or NMP.

Useful concentrations of NMP in the compositions are from about 25%(w/w) to about 44% (w/w). In one embodiment, the concentration of NMP isfrom about 28% (w/w) to about 42% (w/w) or from about 31% (w/w) to about39% (w/w). In still other embodiments, the concentration of NMP is from33% (w/w) to about 37% (w/w). In yet another embodiment, NMP is presentat a concentration of about 35% (w/w). From the level of NMP present orintended, a corresponding amount of diluent can be calculated.

The compositions contain an amount of neutral oil that minimizesfipronil sulfone formation in the formulation comprising fipronil andNMP. The amount of the neutral oil used in the formulations may varyslightly as it is used in an amount to complete the formulation (QS).Useful concentrations of neutral oil(s) in the compositions of theinvention include, but are not limited to, about 11% (w/w) to about 18%(w/w). In some embodiments, concentrations of neutral oil(s) includefrom about 12% (w/w) to about 17% (w/w) and from about 13% (w/w) toabout 16% (w/w). In other embodiments, the neutral oil(s) are present ina concentration of from about 12% (w/w) to about 14% (w/w), about 13%(w/w) to about 15% (w/w). In another embodiment, the neutral oil ispresent at a concentration of about 14% (w/w).

When present in preferred embodiments, an antioxidant is beneficiallypresent at a concentration of not more than about 0.25% (w/w), althoughhigher concentrations could be added. In some embodiments, theconcentration is not more than about: 0.2% (w/w); 0.19% (w/w); 0.18%(w/w); 0.17% (w/w); 0.16% (w/w); 0.15% (w/w); 0.14% (w/w); 0.13% (w/w);0.12% (w/w) or 0.11% (w/w). In another embodiment, if present, theantioxidant is present at a concentration of not more than about 0.1%(w/w). Suitable inorganic antioxidants are, for example, the sulphitesand bisulphites, in particular sodium bisulphite. In one embodiment, theantioxidant is a phenolic antioxidant, such as anisole, butylatedhydroxytoluene and butylated hydroxyanisole, and their mixtures with oneanother. In other embodiments, the antioxidizing agents are thoseconventional in the art and include, for example, ascorbic acid, sodiummetabisulphite, propyl gallate, sodium thiosulphate or a mixture of notmore than two of them. When used in formulations containing NMP and adiluent that does not significantly add peroxide species to theformulation, the amount of antioxidant can be beneficially keptsubstantially below the amount required in formulations that do notcontain the beneficial solvent systems of the invention.

In other embodiments, the present subject matter is directed to apesticidal composition that exhibits surprising and unexpectedrepellency against parasites, including stable flies, comprising fromabout 2% (w/w) to about 15% (w/w) fipronil; from about 30% (w/w) toabout 55% (w/w) permethrin; a glycol, glycol ether, glycol ester orfatty acid ester and N-methyl pyrrolidone, wherein said glycol, glycolether, glycol ester, or fatty acid ester and N-methyl pyrrolidone arepresent in a weight:weight ratio of about 1:1.8 to about 1:2.8, about1:2.0 to about 1:3.5, about 1:2.0 to about 1:3.0, about 1:2.5 to about1:3.5, or about 1:2.5 to about 1:3.0; and optionally an antioxidant.

In another embodiment, the present subject matter is directed to apesticidal composition comprising at least about 5% (w/w) fipronil; atleast about 40% (w/w) permethrin; a glycol, glycol ether, glycol esteror fatty acid ester and N-methyl pyrrolidone, wherein said glycol,glycol ether, glycol ester or fatty acid ester and N-methyl pyrrolidoneare present in a weight:weight ratio of about 1:2.0 to about 1:3.5,about 1:2.0 to about 1:3.0, about 1:2.5 to about 1:3.5, or about 1:2.5to about 1:3.0; and optionally an antioxidant, wherein the compositionprovides unexpected enhanced repellency against parasitic flies,including stable flies.

In another embodiment, the present subject matter is directed to apesticidal composition comprising about 5% (w/w) to about 15% (w/w)fipronil; about 40% (w/w) to about 55% (w/w) permethrin; a glycol,glycol ether, glycol ester or fatty acid ester and N-methyl pyrrolidone,wherein said glycol, glycol ether, glycol ester or fatty acid ester andN-methyl pyrrolidone are present in a ratio of about 1:2.0 to about1:3.5; and optionally an antioxidant.

In another embodiment, the present subject matter is directed to apesticidal composition comprising about 5% (w/w) to about 10% (w/w)fipronil; about 40% (w/w) to about 50% (w/w) permethrin; a glycol,glycol ether, glycol ester or fatty acid ester and N-methyl pyrrolidone,wherein said glycol, glycol ether, glycol ester or fatty acid ester andN-methyl pyrrolidone are present in a ratio of about 1:2.0 to about1:3.5; and optionally an antioxidant.

In another embodiment, the present subject matter is directed to apesticidal composition comprising about 6% (w/w) (w/w) fipronil; about45% (w/w) permethrin; a glycol, glycol ether, glycol ester or fatty acidester and N-methyl pyrrolidone, wherein said glycol, glycol ether,glycol ester or fatty acid ester and N-methyl pyrrolidone are present ina ratio of about 1:2.0 to about 1:3.5; and optionally an antioxidant.

Useful amounts of permethrin and fipronil in these formulations are asdescribed above. In some embodiments, the formulations described aboveprovide unexpected enhanced repellency against parasitic flies,including stable flies, and superior insecticidal efficacy.

In these embodiments, the antioxidant may be present at concentrationsof from about 0.005 to about 1% (w/w) or from about 0.01 to about 0.05%(w/w). In some embodiments, the antioxidant may be present in aconcentration of about 0.01 to about 1% by weight, or from about 0.05%to about 0.5% by weight. In another embodiment, the antioxidant ispresent in a concentration of from about 0.075 to about 0.2% by weight.Amounts lower than 0.2% by weight are useful depending on the peroxidelevels in the diluent that is selected or the NMP.

Glycols useful in the compositions include of diethylene glycol,polyethylene glycol (PEG, including all low molecular weight to highmolecular weight PEG), propylene glycol and polypropylene glycol. Glycolethers include, but are not limited to, methyl diglycol, ethyl diglycol,propyl diglycol, butyl diglycol, methyl glycol, ethyl glycol, ethyleneglycol monomethyl ether, diethylene glycol monomethyl ether, diethyleneglycol monoethyl ether, propylene glycol monomethyl ether andtetrahydrofurfuryl alcohol polyethylene glycol ether (tetraglycol orglycofurol). In one embodiment, the glycol ether is diethylene glycolmonoethyl ether. Glycol esters that may be used in the formulationsinclude carboxylic acid esters of glycols or glycol ethers including,but are not limited to, acetates of glycols and glycol ethers such asethylene glycol monoethyl ether acetate (ethyl CELLOSOLVE™ acetate),ethyleneglycol monobutyl ether acetate (butyl CELLOSOLVE™ acetate),ethyleneglycol monomethyl ether acetate, and the like.

In the embodiment described above comprising a glycol, glycol ether,glycol ester or fatty acid ester and NMP, concentrations of NMP in thecompositions of the invention may be from about 25% (w/w) to about 44%(w/w). In other embodiments, NMP may be present in a concentration ofabout 28% (w/w) to about 42% (w/w) or from about 31% (w/w) to about 39%(w/w). In still other embodiments, NMP may be present at a concentrationof about 33% (w/w) to about 37% (w/w). In another embodiment, NMP may bepresent at a concentration of about 35% (w/w). From the level of NMPpresent or intended, a corresponding amount of diluent can becalculated.

The glycol, glycol ether, glycol ester or fatty acid ester and N-methylpyrrolidone components can be quantified in terms of the relativeamounts of each. Useful weight:weight ratios of glycol, glycol ether,glycol ester or fatty acid ester to NMP include a ratio of from 1:2.0 toabout 1:3.5. In other embodiments, the ratios include from about 1:1.18to about 1:1.28, about 1:2.0 to about 1:3.0; about 1:1.9 to about 1:2.7;about 1:2.0 to about 1:2.6; and about 1:2.5 to about 1:3.0, glycol,glycol ether or fatty acid ester to NMP. In still other embodiments, theweight:weight ratio of glycol, glycol ether or fatty acid ester to NMPwill be 1:1.9, 1:2, 1:2.1, 1:2.2, 1:2.4, 1:2.5, 1:2.6, 1:2.7, 1:2.8,1:2.9, 1:3.0, 1:3.1, 1:3.2, 1:3.3, 1:3.4 or 1:3.5. In other embodimentsof the invention, the weight:weight ratio of glycol, glycol ether,glycol ester or fatty acid ester to NMP will be from about 1:2.5 toabout 1:3.0 or about 1:2.5 to about 1:3.5. These ratios are relativeamounts, but are suitably based on a useful amount of glycol, glycolether or fatty acid ester, or NMP.

In certain embodiments, the compositions contain an amount of glycol,glycol ether, glycol ester or fatty acid ester that minimizes fipronilsulfone formation in the formulation comprising fipronil and NMP.Typically, the amount of glycol, glycol ether, glycol ester or fattyacid ester used in the formulations will an amount sufficient tocomplete the formulation (QS) and will thus vary slightly. Usefulconcentrations of glycol, glycol ether, glycol ester or fatty acid esterinclude from about 11% (w/w) to about 18% (w/w). In other embodiments,the concentration of a glycol, glycol ether, glycol ester or fatty acidester are from about 12% (w/w) to about 17% (w/w) or from about 13%(w/w) to about 16% (w/w). In yet other embodiments of the invention, theconcentration of the glycol, glycol ether or fatty acid ester are fromabout 12% (w/w) to about 14% (w/w) or from about 13% (w/w) to about 15%(w/w). In another embodiment, the glycol, glycol ether or fatty acidester is present at a concentration of about 14% (w/w).

In all embodiments, other solvents and/or diluents may be used in theformulations of the invention. In one embodiment, long-chain alkylamides may also be used so long as they provide the necessary solubilityand physical and chemical stability to maintain the desiredconcentrations of fipronil and permethrin before and after application.These include systems which employ glycerol formal, decanamides andoctanamides, such as N,N-dimethyldecanamide (DMDA). Glycerol formal is amixture of 5-hydroxy-1,3-dioxane and 4-hydroxymethyl-1,3-dioxolane(60:40). In one embodiment, the glycerol formal that is added to aformulation of the invention is stabilized glycerol formal. Stabilizedglycerol formal typically contains 0.02% disodium EDTA, 0.02% N-propylgallate, and 0.01% thiopropionic acid. However, in certain embodimentsfor topical applications, the compositions will include neutral oils asthe diluent.

Additional Active Agents

In one embodiment, the invention provides topical compositionscomprising fipronil and permethrin in combination with one or moreadditional active agents. In some embodiments, the additional activeagents combined with fipronil and permethrin may include, but are notlimited to, acaricides, anthelmintics, insecticides and otherparasiticides of various classes presented herein.

In another embodiment, the topical compositions may also includeveterinary therapeutic agents. Veterinary pharmaceutical agents that maybe included in the compositions of the invention are well-known in theart (see e.g. Plumb' Veterinary Drug Handbook, 5^(th) Edition, ed.Donald C. Plumb, Blackwell Publishing, (2005) or The Merck VeterinaryManual, 9^(th) Edition, (January 2005)) and include but are not limitedto acarbose, acepromazine maleate, acetaminophen, acetazolamide,acetazolamide sodium, acetic acid, acetohydroxamic acid, acetylcysteine,acitretin, acyclovir, albendazole, albuterol sulfate, alfentanil,allopurinol, alprazolam, altrenogest, amantadine, amikacin sulfate,aminocaproic acid, aminopentamide hydrogen sulfate,aminophylline/theophylline, amiodarone, amitriptyline, amlodipinebesylate, ammonium chloride, ammonium molybdenate, amoxicillin,clavulanate potassium, amphotericin B desoxycholate, amphotericin Blipid-based, ampicillin, amprolium, antacids (oral), antivenin,apomorphione, apramycin sulfate, ascorbic acid, asparaginase, aspiring,atenolol, atipamezole, atracurium besylate, atropine sulfate, aurnofin,aurothioglucose, azaperone, azathioprine, azithromycin, baclofen,barbituates, benazepril, betamethasone, bethanechol chloride, bisacodyl,bismuth subsalicylate, bleomycin sulfate, boldenone undecylenate,bromides, bromocriptine mesylate, budenoside, buprenorphine, buspirone,busulfan, butorphanol tartrate, cabergoline, calcitonin salmon,calcitrol, calcium salts, captopril, carbenicillin indanyl sodium,carbimazole, carboplatin, carnitine, carprofen, carvedilol, cefadroxil,cefazolin sodium, cefixime, clorsulon, cefoperazone sodium, cefotaximesodium, cefotetan disodium, cefoxitin sodium, cefpodoxime proxetil,ceftazidime, ceftiofur sodium, ceftiofur, ceftiaxone sodium, cephalexin,cephalosporins, cephapirin, charcoal (activated), chlorambucil,chloramphenicol, chlordiazepoxide, chlordiazepoxide+/−clidinium bromide,chlorothiazide, chlorpheniramine maleate, chlorpromazine,chlorpropamide, chlortetracycline, chorionic gonadotropin (HCG),chromium, cimetidine, ciprofloxacin, cisapride, cisplatin, citratesalts, clarithromycin, clemastine fumarate, clenbuterol, clindamycin,clofazimine, clomipramine, claonazepam, clonidine, cloprostenol sodium,clorazepate dipotassium, clorsulon, cloxacillin, codeine phosphate,colchicine, corticotropin (ACTH), cosyntropin, cyclophosphamide,cyclosporine, cyproheptadine, cytarabine, dacarbazine,dactinomycin/actinomycin D, dalteparin sodium, danazol, dantrolenesodium, dapsone, decoquinate, deferoxamine mesylate, deracoxib,deslorelin acetate, desmopressin acetate, desoxycorticosterone pivalate,detomidine, dexamethasone, dexpanthenol, dexraazoxane, dextran,diazepam, diazoxide (oral), dichlorphenamide, diclofenac sodium,dicloxacillin, diethylcarbamazine citrate, diethylstilbestrol (DES),difloxacin, digoxin, dihydrotachysterol (DHT), diltiazem,dimenhydrinate, dimercaprol/BAL, dimethyl sulfoxide, dinoprosttromethamine, diphenylhydramine, disopyramide phosphate, dobutamine,docusate/DSS, dolasetron mesylate, domperidone, dopamine, doramectin,doxapram, doxepin, doxorubicin, doxycycline, edetate calciumdisodium.calcium EDTA, edrophonium chloride, enalapril/enalaprilat,enoxaparin sodium, enrofloxacin, ephedrine sulfate, epinephrine,epoetin/erythropoietin, eprinomectin, epsiprantel, erythromycin,esmolol, estradiol cypionate, ethacrynic acid/ethacrynate sodium,ethanol (alcohol), etidronate sodium, etodolac, etomidate, euthanasiaagents w/pentobarbital, famotidine, fatty acids (essential/omega),felbamate, fentanyl, ferrous sulfate, filgrastim, finasteride, fipronil,florfenicol, fluconazole, flucytosine, fludrocortisone acetate,flumazenil, flumethasone, flunixin meglumine, fluorouracil (5-FU),fluoxetine, fluticasone propionate, fluvoxamine maleate, fomepizole(4-MP), furazolidone, furosemide, gabapentin, gemcitabine, gentamicinsulfate, glimepiride, glipizide, glucagon, glucocorticoid agents,glucosamine/chondroitin sulfate, glutamine, glyburide, glycerine (oral),glycopyrrolate, gonadorelin, grisseofulvin, guaifenesin, halothane,hemoglobin glutamer-200 (OXYGLOBIN®®), heparin, hetastarch, hyaluronatesodium, hydrazaline, hydrochlorothiazide, hydrocodone bitartrate,hydrocortisone, hydromorphone, hydroxyurea, hydroxyzine, ifosfamide,imidacloprid, imidocarb dipropinate, impenem-cilastatin sodium,imipramine, inamrinone lactate, insulin, interferon alfa-2a (humanrecombinant), iodide (sodium/potassium), ipecac (syrup), ipodate sodium,iron dextran, isoflurane, isoproterenol, isotretinoin, isoxsuprine,itraconazole, ivermectin, kaolin/pectin, ketamine, ketoconazole,ketoprofen, ketorolac tromethamine, lactulose, leuprolide, levamisole,levetiracetam, levothyroxine sodium, lidocaine, lincomycin, liothyroninesodium, lisinopril, lomustine (CCNU), lufenuron, lysine, magnesium,mannitol, marbofloxacin, mechlorethamine, meclizine, meclofenamic acid,medetomidine, medium chain triglycerides, medroxyprogesterone acetate,megestrol acetate, melarsomine, melatonin, meloxican, melphalan,meperidine, mercaptopurine, meropenem, metformin, methadone,methazolamide, methenamine mandelate/hippurate, methimazole, methionine,methocarbamol, methohexital sodium, methotrexate, methoxyflurane,methylene blue, methylphenidate, methylprednisolone, metoclopramide,metoprolol, metronidaxole, mexiletine, mibolerlone, midazolam milbemycinoxime, mineral oil, minocycline, misoprostol, mitotane, mitoxantrone,morphine sulfate, moxidectin, naloxone, mandrolone decanoate, naproxen,narcotic (opiate) agonist analgesics, neomycin sulfate, neostigmine,niacinamide, nitazoxanide, nitenpyram, nitrofurantoin, nitroglycerin,nitroprusside sodium, nizatidine, novobiocin sodium, nystatin,octreotide acetate, olsalazine sodium, omeprozole, ondansetron, opiateantidiarrheals, orbifloxacin, oxacillin sodium, oxazepam, oxibutyninchloride, oxymorphone, oxytretracycline, oxytocin, pamidronate disodium,pancreplipase, pancuronium bromide, paromomycin sulfate, parozetine,pencillamine, general information penicillins, penicillin G, penicillinV potassium, pentazocine, pentobarbital sodium, pentosan polysulfatesodium, pentoxifylline, pergolide mesylate, phenobarbital,phenoxybenzamine, pheylbutazone, phenylephrine, phenypropanolamine,phenytoin sodium, pheromones, parenteral phosphate, phytonadione/vitaminK−1, pimobendan, piperazine, pirlimycin, piroxicam, polysulfatedglycosaminoglycan, ponazuril, potassium chloride, pralidoxime chloride,prazosin, prednisolone/prednisone, primidone, procainamide,procarbazine, prochlorperazine, propantheline bromide, propionibacteriumacnes injection, propofol, propranolol, protamine sulfate,pseudoephedrine, psyllium hydrophilic mucilloid, pyridostigmine bromide,pyrilamine maleate, pyrimethamine, quinacrine, quinidine, ranitidine,rifampin, s-adenosyl-methionine (SAMe), saline/hyperosmotic laxative,selamectin, selegiline/l-deprenyl, sertraline, sevelamer, sevoflurane,silymarin/milk thistle, sodium bicarbonate, sodium polystyrenesulfonate, sodium stibogluconate, sodium sulfate, sodum thiosulfate,somatotropin, sotalol, spectinomycin, spironolactone, stanozolol,streptokinase, streptozocin, succimer, succinylcholine chloride,sucralfate, sufentanil citrate, sulfachlorpyridazine sodium,sulfadiazine/trimethroprim, sulfamethoxazole/trimethoprim,sulfadimentoxine, sulfadimethoxine/ormetoprim, sulfasalazine, taurine,tepoxaline, terbinafline, terbutaline sulfate, testosterone,tetracycline, thiacetarsamide sodium, thiamine, thioguanine, thiopentalsodium, thiotepa, thyrotropin, tiamulin, ticarcilin disodium,tiletamine/zolazepam, tilmocsin, tiopronin, tobramycin sulfate,tocainide, tolazoline, telfenamic acid, topiramate, tramadol,trimcinolone acetonide, trientine, trilostane, trimepraxine tartratew/prednisolone, tripelennamine, tylosin, urdosiol, valproic acid,vanadium, vancomycin, vasopressin, vecuronium bromide, verapamil,vinblastine sulfate, vincristine sulfate, vitamin E/selenium, warfarinsodium, xylazine, yohimbine, zafirlukast, zidovudine (AZT), zincacetate/zinc sulfate, zonisamide and mixtures thereof.

In one embodiment of the invention, additional arylpyrazole compoundssuch as phenylpyrazoles may be included in the topical compositions ofthe invention. The arylpyrazoles are known in the art and are suitablefor combination with the isoxazoline compounds in the soft chewablecompositions of the invention. Examples of such arylpyrazole compoundsinclude but are not limited to those described in U.S. Pat. Nos.6,001,384; 6,010,710; 6,083,519; 6,096,329; 6,174,540; 6,685,954,6,998,131 and 7,759,381 (all of which are incorporated herein byreference).

In another embodiment of the invention, one or more macrocyclic lactonesor lactams, which act as an acaricide, an anthelmintic agent and/or aninsecticide, can be included in the compositions of the invention. Forthe avoidance of doubt, the term “macrocyclic lactone” as used hereinincludes both naturally occurring and synthetic or semi-syntheticavermectin and milbemycin compounds.

The macrocyclic lactones that may be used in the compositions of theinvention include, but are not limited to, the naturally producedavermectins (e.g. including the components designated as A₁a, A₁b, A₂a,A₂b, B₁a, B₁b, B₂a and B₂b) and milbemycin compounds, semisyntheticavermectins and milbemycins, avermectin monosaccharide compounds andavermectin aglycone compounds. Examples of macrocyclic lactone compoundsthat may be used in the compositions include, but are not limited to,abamectin, dimadectin, doramectin, emamectin, eprinomectin, ivermectin,latidectin, lepimectin, selamectin, ML-1,694,554 and milbemycinsincluding, but not limited to, milbemectin, milbemycin D, milbemycin A₃,milbemycin A₄, milbemycin oxime, moxidectin and nemadectin. Alsoincluded are the 5-oxo and 5-oxime derivatives of said avermectins andmilbemycins.

The macrocyclic lactone compounds are known in the art and can easily beobtained commercially or through synthesis techniques known in the art.Reference is made to the widely available technical and commercialliterature. For avermectins, ivermectin and abamectin, reference may bemade, for example, to the work “Ivermectin and Abamectin”, 1989, by M.H. Fischer and H. Mrozik, William C. Campbell, published by SpringerVerlag., or Albers-Schönberg et al. (1981), “Avermectins StructureDetermination”, J. Am. Chem. Soc., 103, 4216-4221. For doramectin,“Veterinary Parasitology”, vol. 49, No. 1, July 1993, 5-15 may beconsulted. For milbemycins, reference may be made, inter alia, to DaviesH. G. et al., 1986, “Avermectins and Milbemycins”, Nat. Prod. Rep., 3,87-121, Mrozik H. et al., 1983, Synthesis of Milbemycins fromAvermectins, Tetrahedron Lett., 24, 5333-5336, U.S. Pat. No. 4,134,973and EP 0 677 054, both incorporated herein by reference.

The structure of the avermectins and milbemycins are closely related,e.g., by sharing a complex 16-membered macrocyclic lactone ring. Thenatural product avermectins are disclosed in U.S. Pat. No. 4,310,519 andthe 22,23-dihydro avermectin compounds are disclosed in U.S. Pat. No.4,199,569. Mention is also made of U.S. Pat. Nos. 4,468,390, 5,824,653,EP 0 007 812 A1, U.K. Patent Specification 1 390 336, EP 0 002 916, andNew Zealand Patent No. 237 086, inter alia. Naturally occurringmilbemycins are described in U.S. Pat. No. 3,950,360 as well as in thevarious references cited in “The Merck Index” 12^(th) ed., S. Budavari,Ed., Merck & Co., Inc. Whitehouse Station, N.J. (1996). Latidectin isdescribed in the “International Nonproprietary Names for PharmaceuticalSubstances (INN)”, WHO Drug Information, vol. 17, no. 4, pp. 263-286,(2003). Semisynthetic derivatives of these classes of compounds are wellknown in the art and are described, for example, in U.S. Pat. Nos.5,077,308, 4,859,657, 4,963,582, 4,855,317, 4,871,719, 4,874,749,4,427,663, 4,310,519, 4,199,569, 5,055,596, 4,973,711, 4,978,677,4,920,148 and EP 0 667 054, all incorporated herein by reference.

In one embodiment, the topical compositions of the invention comprise aneffective amount of at least one of abamectin, dimadectin, doramectin,emamectin, eprinomectin, ivermectin, latidectin, lepimectin, selamectin,milbemectin, milbemycin D, milbemycin A₃, milbemycin A₄, milbemycinoxime, moxidectin or nemadectin, or a combination thereof. In anotherembodiment, the topical veterinary compositions of the invention maycomprise an effective amount of at least one of abamectin, emamectin,eprinomectin, ivermectin, doramectin or selamectin, or a combinationthereof. In still another embodiment, the topical compositions of theinvention may comprise an effective amount of at least one ofivermectin, milbemectin, milbemycin oxime or moxidectin, or acombination thereof.

In another embodiment of the invention, the topical compositioncomprising fipronil and permethrin may include a class of active agentsknown as insect growth regulators (IGRs). Compounds belonging to thisgroup are well known to the practitioner and represent a wide range ofdifferent chemical classes. These compounds all act by interfering withthe development or growth of the insect pests. Insect growth regulatorsare described, for example, in U.S. Pat. Nos. 3,748,356, 3,818,047,4,225,598, 4,798,837, 4,751,225, EP 0 179 022 or U.K. 2 140 010 as wellas U.S. Pat. Nos. 6,096,329 and 6,685,954 (all incorporated herein byreference).

In one embodiment the compositions of the invention may include an IGRcompound that mimics juvenile hormone or that modulates levels ofjuvenile hormones in insects. Examples of juvenile hormone mimicsinclude azadirachtin, diofenolan, fenoxycarb, hydroprene, kinoprene,methoprene, pyriproxyfen, tetrahydroazadirachtin and4-chloro-2(2-chloro-2-methyl-propyl)-5-(6-iodo-3-pyridylmethoxy)pyridazine-3(2H)-one.

In another embodiment, the compositions of the invention include an IGRcompound that is a chitin synthesis inhibitor. Chitin synthesisinhibitors include chlorofluazuron, cyromazine, diflubenzuron,fluazuron, flucycloxuron, flufenoxuron, hexaflumoron, lufenuron,tebufenozide, teflubenzuron, triflumoron,1-(2,6-difluorobenzoyl)-3-(2-fluoro-4-(trifluoromethyl)phenylurea,1-(2,6-difluoro-benzoyl)-3-(2-fluoro-4-(1,1,2,2-tetrafluoroethoxy)-phenylureaand 1-(2,6-difluorobenzoyl)-3-(2-fluoro-4-trifluoromethyl)phenylurea.

In some embodiments, the compositions of the invention may include oneor more antinematodal agents including, but not limited to, activeagents in the benzimidazoles, imidazothiazoles, tetrahydropyrimidinesand the organophosphate class of compounds. In some embodiments,benzimidazoles including, but not limited to, thiabendazole,cambendazole, parbendazole, oxibendazole, mebendazole, flubendazole,fenbendazole, oxfendazole, albendazole, cyclobendazole, febantel,thiophanate and its o,o-dimethyl analogue may be included in thecompositions.

In other embodiments, the compositions may include an imidazothiazolecompounds including, but not limited to, tetramisole, levamisole andbutamisole.

In still other embodiments, the compositions of the invention mayinclude tetrahydropyrimidine active agents including, but not limitedto, pyrantel, oxantel, and morantel.

Suitable organophosphate active agents include, but are not limited to,coumaphos, trichlorfon, haloxon, naftalofos and dichlorvos, heptenophos,mevinphos, monocrotophos, TEPP, and tetrachlorvinphos.

In other embodiments, the compositions may include the antinematodalcompounds phenothiazine, piperazine as the neutral compound and invarious salt forms, diethylcarbamazine, phenols such as disophenol,arsenicals such as arsenamide, ethanolamines such as bephenium, theniumclosylate, and methyridine; cyanine dyes including pyrvinium chloride,pyrvinium pamoate and dithiazanine iodide; isothiocyanates includingbitoscanate, suramin sodium, phthalofyne, and various natural productsincluding, but not limited to, hygromycin B, α-santonin and kainic acid.

In other embodiments, the compositions of the invention may includeantitrematodal agents. Suitable antitrematodal agents include, but arenot limited to, the miracils such as miracil D and mirasan;praziquantel, clonazepam and its 3-methyl derivative, oltipraz,lucanthone, hycanthone, oxamniquine, amoscanate, niridazole, nitroxynil,various bisphenol compounds known in the art including hexachlorophene,bithionol, bithionol sulfoxide and menichlopholan; varioussalicylanilide compounds including tribromsalan, oxyclozanide,clioxanide, rafoxanide, nitroxynil, brotianide, bromoxanide andclosantel; triclabendazole, diamfenetide, clorsulon, hetolin andemetine.

Anticestodal compounds may also be advantageously used in thecompositions of the invention including, but not limited to, arecolinein various salt forms, bunamidine, niclosamide, nitroscanate,paromomycin, paromomycin II, praziquantel and epsiprantel.

In yet other embodiments, the compositions of the invention may includeother active agents that are effective against arthropod parasites.Suitable active agents include, but are not limited to, bromocyclen,chlordane, DDT, endosulfan, lindane, methoxychlor, toxaphene, bromophos,bromophos-ethyl, carbophenothion, chlorfenvinphos, chlorpyrifos,crotoxyphos, cythioate, diazinon, dichlorenthion, diemthoate,dioxathion, ethion, famphur, fenitrothion, fenthion, fospirate,iodofenphos, malathion, naled, phosalone, phosmet, phoxim, propetamphos,ronnel, stirofos, allethrin, cyhalothrin, cypermethrin, deltamethrin,fenvalerate, flucythrinate, permethrin, phenothrin, pyrethrins,resmethrin, benzyl benzoate, carbon disulfide, crotamiton,diflubenzuron, diphenylamine, disulfiram, isobornyl thiocyanato acetate,methoprene, monosulfiram, pirenonylbutoxide, rotenone, triphenyltinacetate, triphenyltin hydroxide, deet, dimethyl phthalate, and thecompounds 1,5a,6,9,9a,9b-hexahydro-4a(4H)-dibenzofurancarboxaldehyde(MGK-11),2-(2-ethylhexyl)-3a,4,7,7a-tetrahydro-4,7-methano-1H-isoindole-1,3(2H)dione(MGK-264), dipropyl-2,5-pyridinedicarboxylate (MGK-326) and2-(octylthio)ethanol (MGK-874).

In another embodiment, an antiparasitic agent that can be included inthe topical veterinary composition can be a biologically active peptideor protein including, but not limited to, depsipeptides, which act atthe neuromuscular junction by stimulating presynaptic receptorsbelonging to the secretin receptor family resulting in the paralysis anddeath of parasites. In one embodiment of the depsipeptide, thedepsipeptide is emodepside (see Wilson et al., Parasitology, January2003, 126(Pt 1):79-86).

In another embodiment, the topical compositions of the invention maycomprise an active agent from the neonicotinoid class of parasiticides.The neonicotinoids bind and inhibit insect specific nicotinicacetylcholine receptors. In one embodiment, the neonicotinoidinsecticidal agent that can be included in the topical compositions ofthe invention is imidacloprid. Agents of this class are described, forexample, in U.S. Pat. No. 4,742,060 or in EP 0 892 060 (bothincorporated herein by reference). In another embodiment, thecompositions of the invention may comprise nitenpyram, another activeagent of the neonicotinoid class of pesticides. The use of nitenpyramfor controlling fleas is described in U.S. Pat. No. 5,750,548, which isincorporated herein by reference in its entirety.

In other certain embodiments of the invention, an insecticidal agentthat can be combined with the compositions of the invention is asemicarbazone, such as metaflumizone.

In another embodiment, the compositions of the invention mayadvantageously include one or more other isoxazoline compounds known inthe art. These active agents, which are very effective againstectoparasites, are described in U.S. Pat. No. 7,964,204 and WO2007/079162; US 2010/0254960 A1, US2011/0159107, US2012/0309620,US2012/0030841, US2010/0069247, WO 2007/125984, WO 2012/086462, U.S.Pat. No. 8,318,757, US 2011/0144349, U.S. Pat. No. 8,053,452; US2010/0137612, US 2010/0254959, US 2011/152081, WO 2012/089623, WO2012/089622, U.S. Pat. No. 8,119,671; U.S. Pat. No. 7,947,715; WO2102/120135, WO 2012/107533, WO 2011/157748, US 2011/0245274, US2011/0245239, US 2012/0232026, US 2012/0077765, US 2012/0035122, US2011/0251247, WO 2011/154433, WO 2011/154434, US 2012/0238517, US2011/0166193, WO 2011/104088, WO 2011/104087, WO 2011/104089, US2012/015946, US 2009/0143410, WO 2007/123855 A2, US 2011/0118212, U.S.Pat. No. 7,951,828 & U.S. Pat. No. 7,662,972, US 2010/0137372 A1, US2010/0179194 A2, US 2011/0086886 A2, US 2011/0059988 A1, US 2010/0179195A1, U.S. Pat. No. 7,897,630, U.S. Pat. No. 7,951,828; WO 2011/075591 andUS 2011/0152312, and U.S. Pat. No. 7,662,972, all of which areincorporated herein by reference in their entirety.

In another embodiment of the invention, nodulisporic acid and itsderivatives may be added to the compositions of the invention. Thesecompounds are used to treat or prevent infections in humans and animalsand are described, for example, in U.S. Pat. Nos. 5,399,582, 5,962,499,6,221,894 and 6,399,786, all of which are hereby incorporated byreference in their entirety. The compositions may include one or more ofthe known nodulisporic acid derivatives in the art, including allstereoisomers, such as those described in the literature cited above.

In another embodiment, anthelmintic compounds of the amino acetonitrileclass (AAD) of compounds such as monepantel (ZOLVIX) and the like may beadded to the compositions of the invention. These compounds aredescribed, for example, in U.S. Pat. No. 7,084,280 to Ducray et al.(incorporated herein by reference); Sager et al., VeterinaryParasitology, 2009, 159, 49-54; Kaminsky et al., Nature vol. 452, 13Mar. 2008, 176-181.

The compositions of the invention may also include aryloazol-2-ylcyanoethylamino compounds such as those described in U.S. Pat. No.8,088,801 and US 2010/0125089 to Soll et al., which is incorporatedherein by reference, and thioamide derivatives of these compounds, asdescribed in U.S. Pat. No. 7,964,621 to Le Hir de Fallois, which is alsoincorporated herein by reference.

The compositions of the invention may also include paraherquamidecompounds and derivatives of these compounds, including derquantel (seeOstlind et al., Research in Veterinary Science, 1990, 48, 260-61; andOstlind et al., Medical and Veterinary Entomology, 1997, 11, 407-408).The paraherquamide family of compounds is a known class of compoundsthat include a spirodioxepino indole core with activity against certainparasites (see Tett. Lett. 1981, 22, 135; J. Antibiotics 1990, 43, 1380,and J. Antibiotics 1991, 44, 492). In addition, the structurally relatedmarcfortine family of compounds, such as marcfortines A-C, are alsoknown and may be combined with the formulations of the invention (see J.Chem. Soc.—Chem. Comm. 1980, 601 and Tet. Lett. 1981, 22, 1977). Furtherreferences to the paraherquamide derivatives can be found, for example,in WO 91/09961, WO 92/22555, WO 97/03988, WO 01/076370, WO 09/004432 andUS 2010/0197624, U.S. Pat. No. 5,703,078 and U.S. Pat. No. 5,750,695,all of which are hereby incorporated by reference in their entirety.

In another embodiment of the invention, the compositions may include aspinosyn active agent produced by the soil actinomyceteSaccharopolyspora spinosa (see, for example Salgado V. L. and Sparks T.C., “The Spinosyns: Chemistry, Biochemistry, Mode of Action, andResistance,” in Comprehensive Molecular Insect Science, vol. 6, pp.137-173, 2005) or a semi-synthetic spinosoid active agent. The spinosynsare typically referred to as factors or components A, B, C, D, E, F, G,H, J, K, L, M, N, O, P, Q, R, S, T, U, V, W, or Y, and any of thesecomponents, or a combination thereof, may be used in the compositions ofthe invention. The spinosyn compound may be a 5,6,5-tricylic ringsystem, fused to a 12-membered macro cyclic lactone, a neutral sugar(rhamnose), and an amino sugar (forosamine). These and other naturalspinosyn compounds, including 21-butenyl spinosyn produced bySaccharopolyspora pagona, which may be used in the compositions of theinvention, may be produced via fermentation by conventional techniquesknown in the art. Other spinosyn compounds that may be used in thecompositions of the invention are disclosed in U.S. Pat. Nos. 5,496,931;5,670,364; 5,591,606; 5,571,901; 5,202,242; 5,767,253; 5,840,861;5,670,486; 5,631,155 and 6,001,981, all incorporated by reference hereinin their entirety. The spinosyn compounds may include, but are notlimited to, spinosyn A, spinosyn D, spinosad, spinetoram, orcombinations thereof. Spinosad is a combination of spinosyn A andspinosyn D, and spinetoram is a combination of 3′-ethoxy-5,6-dihydrospinosyn J and 3′-ethoxy spinosyn L.

In general, the additional active agent is included in the dosage unitsof the invention in an amount of between about 0.1 μg and about 1000 mg.Typically, the active agent may be included in an amount of about 10 μgto about 500 mg, about 10 μg to about 400 mg, about 1 mg to about 300mg, about 10 mg to about 200 mg or about 10 mg to about 100 mg. Moretypically the active agent will be present in an amount of about 5 mg toabout 50 mg in the compositions of the invention.

The concentration of the additional active agent(s) in the topicalcompositions of the invention will typically be from about 0.01% toabout 30% (w/w) depending on the potency of the active agent. In certainembodiments for very potent active agents including, but not limited toa macrocyclic lactone active agent, the concentration of the activeagent will typically be from about 0.01% to about 10% (w/w), from about0.01 to about 1% (w/w), from about 0.01% to about 0.5% (w/w), from about0.1% to about 0.5% (w/w) or from about 0.01% to about 0.1% (w/w). Inother embodiments, the concentration of the active agent will typicallybe from about 0.1% to about 2% (w/w) or about 0.1% to about 1% (w/w).

In other embodiments, the additional active agent(s) will typically bepresent at higher concentrations to achieve the desired efficacy. Insome embodiments, the active agent will be present in a concentration ofabout 1% to about 30% (w/w), about 1% to about 20% (w/w) or about 1% toabout 15% (w/w). In still other embodiments, the active agent will bepresent in a concentration of about 5% to about 20% (w/w) or about 5% toabout 15% (w/w) in the composition.

In various embodiments of the invention, an additional active agent maybe included in the composition to deliver a dose of about 0.001 mg/kg toabout 50 mg/kg or about 0.5 mg/kg to about 50 mg/kg of body weight ofthe animal. In other embodiments, the active agent will typically bepresent in an amount sufficient to deliver a dose of about 0.05 mg/kg toabout 30 mg/kg, about 0.1 mg/kg to about 20 mg/kg. In other embodiments,the active agent will be present in an amount sufficient to deliver adose of about 0.1 mg/kg to about 10 mg/kg, about 0.1 mg/kg to about 1mg/kg or about 0.5 mg/kg to about 50 mg/kg per body weight of theanimal.

In certain embodiments of the invention where the additional activeagent is a very potent compound such as a macrocyclic lactone or otherpotent compounds, the active agent will be present in a concentration toprovide a dose of about 0.001 mg/kg to about 5 mg/kg, about 0.001 mg/kgto about 0.1 mg/kg or about 0.001 mg/kg to about 0.01 mg/kg. In stillother embodiments, the active agent is present in an amount sufficientto deliver a dose of about 0.01 mg/kg to about 2 mg/kg or about 0.1mg/kg to about 1 mg/kg per body weight of the animal. In still otherembodiments, the additional active agent may be present in an amount todeliver a dose of about 1 μg/kg to about 200 μg/kg or about 0.1 mg/kg toabout 1 mg/kg of weight of animal.

Methods and Uses

In an embodiment, the present subject matter is directed to a method oftreating and/or controlling a parasite infestation in an animal. Themethod comprises administering an effective amount of a formulationdescribed herein to the animal. Surprisingly, it has been found anddisclosed herein that at particular concentrations, fipronil andpermethrin have surprisingly enhanced repellent activity against stablefly. In one embodiment, the formulations of the invention having acombination of fipronil and permethrin were unexpectedly found to have asignificantly higher repellent efficacy against stable flies compared toa formulation containing permethrin alone at the same concentrationadministered to deliver the same dose. The observed enhanced repellentefficacy is surprising because fipronil, although a very potentinsecticidal agent, is not known to have repellent efficacy againstectoparasites. Accordingly, one method of the invention is directed tocombatting pest infestations of parasitic flies including stable fly(Stomoxys calcitrans) and horn fly (Haematobia irritans), among otherfly species.

In other embodiments, the uses and methods of the invention are fortreating and/or preventing parasite infestations and/or infections frompests including those from the order of the Anoplura, for example,Haematopinus spp., Linognathus spp., Solenopotes spp., Pediculus spp.,Pthirus spp.; from the order of the Mallophaga, for example, Trimenoponspp., Menopon spp., Eomenacanthus spp., Menacanthus spp., Trichodectesspp., Felicola spp., Damalinea spp., Bovicola spp.; from the order ofthe Diptera, suborder Brachycera, for example, Chrysops spp., Tabanusspp., Musca spp., Hydrotaea spp., Muscina spp., Haematobosca spp.,Haematobia spp., Stomoxys spp., Fannia spp., Glossina spp., Luciliaspp., Calliphora spp., Auchmeromyia spp., Cordylobia spp., Cochliomyiaspp., Chrysomyia spp., Sarcophaga spp., Wohlfahrtia spp., Gasterophilusspp., Oedemagena spp., Hypoderma spp., Oestrus spp., Rhinoestrus spp.,Melophagus spp., Hippobosca spp.; from the order of the Diptera,suborder Nematocera, for example, Culex spp., Aedes spp., Anophelesspp., Culicoides spp., Phlebotomus spp., Simulium spp.; from the orderof the Siphonaptera, for example, Ctenocephalides spp., Echidnophagaspp., Ceratophyllus spp., Pulex spp.; from the order of theMetastigmata, for example, Hyalomma spp., Rhipicephalus spp., Boophilusspp., Amblyomma spp., Haemaphysalis spp., Dermacentor spp., Ixodes spp.,Argas spp., Otobius spp.; from the order of the Mesostigmata, forexample, Dermanyssus spp., Ornithonyssus spp., Pneumonyssus spp.; fromthe order of the Prostigmata, for example, Cheyletiella spp.,Psorergates spp., Myobia spp., Demodex spp., Neotrombicula spp.; fromthe order of the Astigmata, for example, Acarus spp., Myocoptes spp.,Psoroptes spp., Chorioptes spp., Otodectes spp., Sarcoptes spp.,Notoedres spp., Knemidocoptes spp., Neoknemidocoptes spp., Cytoditesspp., Laminosioptes spp.; and fleas (Siphonaptera, for example,Ctenocephalides spp., Echidnophaga spp., Ceratophyllus spp., Pulexspp.), ticks (Hyalomma spp., Rhipicephalus spp., Boophilus spp.,Amblyomma spp., Haemaphysalis spp., Dermacentor spp., Ixodes spp., Argasspp., Ornithodorus spp., Otobius spp.) and in particular the Dipteramentioned above (Chrysops spp., Tabanus spp., Musca spp., Hydrotaeaspp., Muscina spp., Haematobosca spp., Haematobia spp., Stomoxys spp.,Fannia spp., Glossina spp., Lucilia spp., Calliphora spp., Auchmeromyiaspp., Cordylobia spp., Cochliomyia spp., Chrysomyia spp., Sarcophagaspp., Gasterophilus spp., Oedemagena spp., Hypoderma spp., Oestrus spp.,Rhinoestrus spp., Melophagus spp., Hippobosca spp.).

In another embodiment, the subject matter disclosed herein is directedto a method for cleaning the coats and the skin of animals by removal ofthe parasites which are present and of their waste and excreta andadministering to the animal a formulation as described herein. Theanimals treated thus exhibit a coat which is more pleasing to the eyeand more pleasant to the touch.

The term “animal” as used herein refers to any mammal or bird. Inparticular, cattle, sheep, horses, pigs, chicken, and dogs would benefitfrom administration of the compositions disclosed herein as they may beinfested by parasites that are effectively controlled by thecompositions. The useful and breeding animals include mammals, such as,for example, cattle, horses, zebras, sheep, pigs, goats, camels, yaks,water buffalo, donkeys, fallow deer, reindeer, rodents, fur-bearinganimals, such as, for example, mink, chinchilla, raccoon, birds, suchas, for example, hens, chickens, geese, turkeys, quails and ducks.Domesticated animals include dogs, and particular emphasis is given tothe treatment of dogs.

Various methods of formulating parasiticidal formulations are known inthe art. These include oral formulations, dietary supplements, powders,skin solutions (pour-on or spot-on), sprays, drenches, baths, showers,jets, powders, greases, shampoos and creams.

Of particular interest are formulations for localized topicalapplications of parasiticidal active agents known in the art as spot-onor pour-on formulations. Each type of formulation has distinctproperties. For example, pour-on solutions comprising 1-N-phenylpyrazolederivatives, such as fipronil, are known in the art and are describedin, for example, U.S. Pat. No. 6,010,710, U.S. Pat. No. 6,413,542, U.S.Pat. No. 6,001,384 and U.S. Pat. No. 6,413,542. Spot-on formulations arewell known techniques for topically delivering certain antiparasiticagents with the required physicochemical properties to a limited area ofthe host. For example, U.S. Pat. No. 5,045,536 describes suchformulations for ectoparasites. Other spot-on formulations include U.S.Pat. No. 6,426,333 and U.S. Pat. No. 6,482,425. All of these referencesare hereby incorporated herein in their entirety.

In one embodiment, the composition is a spot-on formulation comprisingfrom about 2% (w/w) to about 10% (w/w) fipronil; from about 30% (w/w) toabout 55% (w/w) permethrin; and a glycol, glycol ether, glycol ester,fatty acid ester or neutral oil and N-methyl pyrrolidone, wherein saidglycol, glycol ether, glycol ester, fatty acid ester or neutral oil andN-methyl pyrrolidone are present in a ratio of from about 1:1.8 to about1:2.8 or about 1:2.0 to about 1:3.5, glycol, glycol ether, glycol ester,fatty acid ester or neutral oil to NMP, and further wherein saidcomposition is a liquid having a volume of from about 0.5 mL to about 10mL. In another embodiment, the volume of the spot-on composition is fromabout 1 mL to about 6 mL.

In another embodiment, the composition is a spot-on formulationcomprising at least about 5% (w/w) fipronil; at least about 40% (w/w)permethrin; and a glycol, glycol ether, glycol ester, fatty acid esteror neutral oil and N-methyl pyrrolidone, wherein said glycol, glycolether, glycol ester, fatty acid ester or neutral oil and N-methylpyrrolidone are present in a weight:weight ratio of from about 1:1.8 toabout 1:2.8 or about 1:2.0 to about 1:3.5, glycol, glycol ether, glycolester, fatty acid ester or neutral oil to NMP, and further wherein saidcomposition is a liquid having a volume of from about 0.5 mL to about 10mL. In another embodiment, the volume of the spot-on composition is fromabout 1 mL to about 6 mL.

In one embodiment, the composition is a spot-on formulation comprisingat least about 5% (w/w) fipronil; at least about 40% (w/w) permethrin;and a glycol, glycol ether, glycol ester, fatty acid ester or neutraloil and N-methyl pyrrolidone, wherein said glycol, glycol ether, glycolester, fatty acid ester or neutral oil and N-methyl pyrrolidone arepresent in a weight:weight ratio of from about 1:2.5 to about 1:3.5 orabout 1:25 to about 1:3.0, glycol, glycol ether, glycol ester, fattyacid ester or neutral oil to NMP, and further wherein said compositionis a liquid having a volume of from about 0.5 mL to about 10 mL. Inanother embodiment, the volume of the spot-on composition is from about1 mL to about 6 mL.

In still another embodiment, the composition is a spot-on formulationcomprising about 5% (w/w) to about 15% (w/w) fipronil; about 40% (w/w)to about 55% (w/w) permethrin; and a glycol, glycol ether, glycol ester,fatty acid ester or neutral oil and N-methyl pyrrolidone, wherein saidglycol, glycol ether, glycol ester, fatty acid ester or neutral oil andN-methyl pyrrolidone are present in a weight:weight ratio of from about1:1.8 to about 1:2.8 or about 1:2.0 to about 1:3.5, glycol, glycolether, glycol ester, fatty acid ester or neutral oil to NMP, and furtherwherein said composition is a liquid having a volume of from about 0.5mL to about 10 mL. In another embodiment, the volume of the spot-oncomposition is from about 1 mL to about 6 mL.

In still another embodiment, the composition is a spot-on formulationcomprising about 5% (w/w) to about 15% (w/w) fipronil; about 40% (w/w)to about 55% (w/w) permethrin; and a glycol, glycol ether, glycol ester,fatty acid ester or neutral oil and N-methyl pyrrolidone, wherein saidglycol, glycol ether, glycol ester, fatty acid ester or neutral oil andN-methyl pyrrolidone are present in a weight:weight ratio of from about1:2.5 to about 1:3.0 or about 1:2.5 to about 1:3.5, glycol, glycolether, glycol ester, fatty acid ester or neutral oil to NMP, and furtherwherein said composition is a liquid having a volume of from about 0.5mL to about 10 mL. In another embodiment, the volume of the spot-oncomposition is from about 1 mL to about 6 mL.

In another embodiment, the composition is a spot-on formulationcomprising about 5% (w/w) to about 10% (w/w) fipronil; about 40% (w/w)to about 50% (w/w) permethrin; and a glycol, glycol ether, glycol ester,fatty acid ester or neutral oil and N-methyl pyrrolidone, wherein saidglycol, glycol ether, glycol ester, fatty acid ester or neutral oil andN-methyl pyrrolidone are present in a weight:weight ratio of from about1:1.8 to about 1:2.8 or about 1:2.0 to about 1:3.5, glycol, glycolether, glycol ester, fatty acid ester or neutral oil to NMP, and furtherwherein said composition is a liquid having a volume of from about 0.5mL to about 10 mL. In another embodiment, the volume of the spot-oncomposition is from about 1 mL to about 6 mL.

In another embodiment, the composition is a spot-on formulationcomprising about 5% (w/w) to about 10% (w/w) fipronil; about 40% (w/w)to about 50% (w/w) permethrin; and a glycol, glycol ether, glycol ester,fatty acid ester or neutral oil and N-methyl pyrrolidone, wherein saidglycol, glycol ether, glycol ester, fatty acid ester or neutral oil andN-methyl pyrrolidone are present in a weight:weight ratio of from about1:2.5 to about 1:3.5 or about 1:2.5 to about 1:3.0, glycol, glycolether, glycol ester, fatty acid ester or neutral oil to NMP, and furtherwherein said composition is a liquid having a volume of from about 0.5mL to about 10 mL. In another embodiment, the volume of the spot-oncomposition is from about 1 mL to about 6 mL.

The topical composition of the invention can be administered in severalways. Administering comprises contacting the coat and/or skin of saidanimal with said composition. In one embodiment, the formulation is aspot-on or pour-on formulation. These formulations are applied to theanimal's back, along the line of the back for a pour-on formulation orat one or two spots on the back for a spot-on product. In oneembodiment, the formulation will be administered to the animal over ahighly localized region of the animal, preferably between the shoulderblades. In another embodiment, this localized region has a surface areaof less than 10 cm², especially between 5 and 10 cm² area. Suchcontacting or application can take place both prophylactically andtherapeutically.

Administration of the formulations may be intermittent in time and maybe administered daily, weekly, biweekly, monthly, bimonthly, quarterly,or even for longer durations of time. The time period between treatmentsdepends upon factors such as the target parasite(s), the degree ofinfestation, the type of mammal or bird being treated and theenvironment where it resides. It is well within the skill level of thepractitioner to determine a specific administration period for aparticular situation. The presently described methods are directed topermanently combating a parasite in an environment in which the animalis subjected to strong parasitic pressure where the administration is ata frequency far below a daily administration in this case. For example,in one embodiment the treatment according to the invention will becarried out monthly on mammals such as dogs.

In certain embodiments, the liquid formulations according to theinvention are suitable for spray application, where the sprayapplication may be carried out, for example, using a pump spray or anaerosol spray (pressurized spray). For specific indications, theformulations may also be used after dilution with water as a dip; inthis case, the formulation should contain emulsifying additives. In oneembodiment, the compositions are applied as pour-on and spot-onformulations. The spot-on application is particularly preferred. Theformulations according to the invention are distinguished by theirexcellent compatibility with customary “single-dose” plastic pipettesand by their storage stability in various climate zones. They have lowviscosity and can be easily applied.

The liquid formulations according to the invention can be prepared bymixing appropriate amounts of the components with one another, using,for example, conventional stirring tanks or other suitable instruments.If required by the ingredients, it is also possible to operate under aprotective atmosphere or with other methods of excluding oxygen. Spot-onformulations may be prepared by dissolving the active ingredients intothe pharmaceutically or veterinary acceptable vehicle. Alternatively,the spot-on formulation can be prepared by encapsulation of the activeingredient to leave a residue of the therapeutic agent on the surface ofthe animal. These formulations will vary with regard to the weight ofthe therapeutic agent in the combination depending on the species ofhost animal to be treated, the severity and type of infection and thebody weight of the host.

In certain embodiments, compositions described herein do not containvisible solids and are clear, but not necessarily water-clear, solutionshaving the described synergistic amounts of fipronil and permethrinfully solubilized therein. The two-component solvent system secures theobjectives of absence of crystallization on the coat and of maintenanceof the cosmetic appearance of the fur, without a tendency towardssticking or towards a sticky appearance, despite the high concentrationof actives in the composition.

In some embodiments the formulations disclosed herein can also comprisesynergists. Synergists in the sense of this application are to beunderstood as meaning compounds which for their part do not have thedesired activity, but which, as mixing partners, increase the activityof the active compounds. Piperonyl butoxide, MGK264, verbutin,S,S,S-tributyl phosphorotrithioate may be mentioned here in an exemplarymanner.

Though not required, the compositions may further include a stabilityenhancer. A “stability enhancer” is a compound that enhances thestability of an active agent compared to the stability of the activeagent in the absence of the stability enhancer. Examples of stabilityenhancers include glycerol formal and polyethylene glycol (e.g., PEG200). Other stability enhancers are well-known in this field. Inparticular embodiments, a small amount of glycerol formal, e.g., 5% orless, can be added. In some embodiments, it may be desirable to add PEG200 to the formulations of the invention to support the stabilityenhancement and solvency function of the glycerol formal.

The invention further provides for titration of the amount of stabilityenhancer that is added to the formulations of the invention. Forexample, in embodiments where the stability enhancer is glycerol formal,the amount of glycerol formal can be titrated, such that an optimalstability of the fipronil and permethrin composition is achieved in theformulation. The amount of stability enhancer present in a formulationof the invention may be low, such as about 5% w/v or less (e.g., 1.5%w/v). In other embodiments, the stability enhancer will be present in anamount of about 5-25% w/v, such as, for example, 15% w/v.

In some embodiments the solutions according to the invention which areoily, in addition to a glycol ether, may comprise a diluent or vehicleand also a solvent (organic solvent) for the active agent(s). However,due to the ability of the two-component solvent system described hereinto solubilize both permethrin and fipronil, the use of additionsolubility enhancers would suitably be minimal.

Further, though not required, if desired, crystallization inhibitors canbe used in the compositions. These include: polyvinylpyrrolidone,polyvinyl alcohols, copolymers of vinyl acetate and of vinylpyrrolidone,polyethylene glycols, benzyl alcohol, mannitol, glycerol, sorbitol orpolyoxyethylenated esters of sorbitan; lecithin or sodiumcarboxymethylcellulose; or acrylic derivatives, such as methacrylatesand others, anionic surfactants, such as alkaline stearates, inparticular sodium, potassium or ammonium stearate; calcium stearate ortriethanolamine stearate; sodium abietate; alkyl sulphates, inparticular sodium lauryl sulphate and sodium cetyl sulphate; sodiumdodecylbenzenesulphonate or sodium dioctyl sulphosuccinate; or fattyacids, in particular those derived from coconut oil, cationicsurfactants, such as water-soluble quaternary ammonium salts of formulaN⁺R′R″R′″R″″Y⁻, in which the R′, R″, R′″ and R″″ radicals are identicalor different optionally hydroxylated hydrocarbon radicals and Y⁻ is ananion of a strong acid, such as halide, sulphate and sulphonate anions;cetyltrimethylammonium bromide is one of the cationic surfactants whichcan be used, amine salts of formula N⁺HR′R″R′″ Y⁻, in which the R′, R″,and R′″ radicals are identical or different optionally hydroxylatedhydrocarbon radicals and Y⁻ is as defined above; octadecylaminehydrochloride is one of the cationic surfactants which can be used,non-ionic surfactants, such as optionally polyoxyethylenated esters ofsorbitan, in particular Polysorbate 80, or polyoxyethylenated alkylethers; polyethylene glycol stearate, polyoxyethylenated derivatives ofcastor oil, polyglycerol esters, polyoxyethylenated fatty alcohols,polyoxyethylenated fatty acids or copolymers of ethylene oxide and ofpropylene oxide, amphoteric surfactants, such as substituted laurylcompounds of betaine, or a mixture of at least two of the compoundslisted above.

A crystallization inhibitor pair can be used. Such pairs include, forexample, the combination of a film-forming agent of polymeric type andof a surface-active agent. These agents will be selected in particularfrom the compounds mentioned above as crystallization inhibitor. Incertain embodiments of the invention, film-forming agents of polymerictype include the various grades of polyvinylpyrrolidone, polyvinylalcohols, polyethylene glycols and copolymers of vinyl acetate and ofvinylpyrrolidone. In one embodiment, surface-active agents include thosemade of non-ionic surfactants, including polyoxyethylenated esters ofsorbitan and polyoxyethylenated derivatives of castor oil, and inparticular the various grades of polysorbate, for example Polysorbate80.

The film-forming agent and the surface-active agent can in particular beincorporated in similar or identical amounts within the limit of thetotal amounts of crystallization inhibitor mentioned elsewhere.

Alternative or additional organic solvents which can be used in theinvention include acetyltributyl citrate, fatty acid esters such as thedimethyl ester, diisobutyl adipate, acetone, acetonitrile, benzylalcohol, butyl diglycol, dimethylacetamide, dimethylformamide,dimethylsulfoxide (DMSO), diethyl sebacate, dimethyl isosorbide,dipropylene glycol n-butyl ether, ethanol, isopropanol, methanol,ethylene glycol monoethyl ether, ethylene glycol monomethyl ether,monomethylacetamide, dipropylene glycol monomethyl ether, liquidpolyoxyethylene glycols, propylene glycol, 2-pyrrolidone, ethyleneglycol; glycol esters including ethylene glycol monoethyl ether acetate,ethylene glycol monobutyl ether acetate, ethylene glycol monomethylether acetate, and the like; and diethyl phthalate, or a mixture of atleast two of these solvents.

In addition, mention may be made in particular of other possibleingredients such as plant oils: soybean oil, groundnut oil, castor oil,corn oil, cotton oil, olive oil, grape seed oil, sunflower oil, etc.;mineral oils such as petrolatum, paraffin, silicone, etc.; aliphatic orcyclic hydrocarbons or alternatively, for example, medium-chain (C8 toC12 in particular) triglycerides.

An emollient and/or spreading and/or film-forming agent may additionallybe added, this agent being selected in particular from:polyvinylpyrrolidone, polyvinyl alcohols, copolymers of vinyl acetateand vinylpyrrolidone, polyethylene glycols, benzyl alcohol, mannitol,glycerol, sorbitol, polyoxyethylenated sorbitan esters; lecithin, sodiumcarboxymethylcellulose, silicone oils, polydiorganosiloxane oils, inparticular polydimethylsiloxane (PDMS) oils, for example thosecontaining silanol functionalities, or a 45V2 oil, anionic surfactantssuch as alkaline stearates, in particular sodium, potassium or ammoniumstearates; calcium stearate, triethanolamine stearate; sodium abietate;alkyl sulphates, in particular sodium lauryl sulphate and sodium cetylsulphate; sodium dodecylbenzenesulphonate, sodiumdioctylsulphosuccinate; fatty acids, in particular those derived fromcoconut oil, cationic surfactants such as water-soluble quaternaryammonium salts of formula N⁺R′R″R′″R″″, Y⁻ in which the radicals R′, R″,R′″ and R″″ are optionally hydroxylated hydrocarbon radicals and Y is ananion of a strong acid such as the halide, sulphate and sulphonateanions; cetyltrimethylammonium bromide is among the cationic surfactantswhich can be used, amine salts of formula N⁺HR″R′″R″″, Y in which theradicals the R″, R′″ and R″″ are optionally hydroxylated hydrocarbonradicals and Y is as defined above; octadecylamine hydrochloride isamong the cationic surfactants which can be used, nonionic surfactantssuch as sorbitan esters, which are optionally polyoxyethylenated, inparticular polysorbate 80, polyoxyethylenated alkyl ethers;polyoxypropylated fatty alcohols such as polyoxypropylene-styrol ether;polyethylene glycol stearate, polyoxyethylenated derivatives of castoroil, polyglycerol esters, polyoxyethylenated fatty alcohols,polyoxyethylenated fatty acids, copolymers of ethylene oxide andpropylene oxide, amphoteric surfactants such as the substituted laurylcompounds of betaine; or a mixture of at least two of these agents. Theemollient may be used in a proportion of from 0.1 to 10%, in particularfrom 0.25 to 5%, by volume.

In some embodiments, the liquid carrier will contain an organic solventand optionally an organic co-solvent. In one embodiment, the organicsolvent for the liquid carrier vehicle will have a dielectric constantof between about 10 and about 35. In another embodiment, the organicsolvent will have a dielectric constant of between about 20 and about30, the content of this solvent in the overall composition representingthe remainder to 100% of the composition. It is well within the skilllevel of the practitioner to select a suitable solvent on the basis ofthese parameters.

In some embodiments, the organic co-solvent for the liquid carriervehicle will have a boiling point of less than about 100° C. In anotherembodiment, the organic co-solvent will have a boiling point of lessthan about 80° C., and will have a dielectric constant of between about10 and about 40. In still another embodiment, the organic co-solventwill have a dielectric constant of between about 20 and about 30. Thisco-solvent can advantageously be present in the composition according toa weight/weight (w/w) ratio with respect to the solvent of between about1/15 and about 1/2. In one embodiment, the co-solvent will be volatilein order to act in particular as drying promoter and is miscible withwater and/or with the solvent. Again, it is well within the skill levelof the practitioner to select a suitable solvent on the basis of theseparameters.

The organic solvent for the liquid carrier includes the commonlyacceptable organic solvents known in the formulation art. These solventsmay be found, for example, in Remington Pharmaceutical Science, 16thEdition (1986). These solvents include, for example, acetone, ethylacetate, methanol, ethanol, isopropanol, dimethylformamide ordichloromethane. Those solvents which are suitable for topicalapplication are preferred.

In another embodiment, the subject matter disclosed herein is directedto a solvent system comprising A) NMP and B) a glycol, glycol ether,glycol ester, fatty acid ester or neutral oil, wherein the N-methylpyrrolidone and glycol, glycol ether, glycol ester, fatty acid ester orneutral oil are present in a weight:weight ratio of from about 1:1.8 toabout 1:2.8, glycol, glycol ether, glycol ester, fatty acid ester orneutral oil to NMP, wherein dissolved in the solvent system are twoactive agents selected from one N-arylpyrazole and one pyrethroid. Inone such embodiment, the present subject matter is directed to aformulation comprising, one pyrethroid in a concentration of from about30% w/w to about 55% w/w; one N-arylpyrazole in a concentration of fromabout 2% w/w to about 10% w/w; and A) glycol, glycol ether, glycolester, fatty acid ester or neutral oil and B) NMP, wherein the glycol,glycol ether, glycol ester, fatty acid ester or neutral oil and NMP arepresent in a weight:weight ratio of from about 1:1.8 to about 1:2.8,glycol, glycol ether, glycol ester, fatty acid ester or neutral oil toNMP.

In another embodiment, the formulation comprises one pyrethroid in anconcentration of at least about 40% w/w; one N-arylpyrazole in aconcentration of at least about 5% w/w; and A) glycol, glycol ether,glycol ester, fatty acid ester or neutral oil and B) NMP, wherein theglycol, glycol ether, glycol ester, fatty acid ester or neutral oil andNMP are present in a weight:weight ratio of from about 1:1.8 to about1:2.8 or about 1:2.0 to about 1:3.5, glycol, glycol ether, glycol ester,fatty acid ester or neutral oil to NMP.

In another embodiment, the formulation comprises one pyrethroid in anconcentration of at least about 40% w/w; one N-arylpyrazole in aconcentration of at least about 5% w/w; and A) glycol, glycol ether,glycol ester, fatty acid ester or neutral oil and B) NMP, wherein theglycol, glycol ether, glycol ester, fatty acid ester or neutral oil andNMP are present in a weight:weight ratio of from about 1:2.0 to about1:3.0, glycol, glycol ether, glycol ester, fatty acid ester or neutraloil to NMP.

In another embodiment, the formulation comprises one pyrethroid in anconcentration of at least about 40% w/w; one N-arylpyrazole in aconcentration of at least about 5% w/w; and A) glycol, glycol ether,glycol ester, fatty acid ester or neutral oil and B) NMP, wherein theglycol, glycol ether, glycol ester, fatty acid ester or neutral oil andNMP are present in a weight:weight ratio of from about 1:2.5 to about1:3.5, glycol, glycol ether, glycol ester, fatty acid ester or neutraloil to NMP.

In another embodiment, the formulation comprises one pyrethroid in anconcentration of at least about 40% w/w; one N-arylpyrazole in aconcentration of at least about 5% w/w; and A) glycol, glycol ether,glycol ester, fatty acid ester or neutral oil and B) NMP, wherein theglycol, glycol ether, glycol ester, fatty acid ester or neutral oil andNMP are present in a ratio of from about 1:2.5 to about 1:3.0, glycol,glycol ether, glycol ester, fatty acid ester or neutral oil to NMP.

In specific aspects of this embodiment, the present subject matter isdirected to a pesticidal composition comprising: from about 2% (w/w) toabout 15% (w/w) N-arylpyrazole; from about 30% (w/w) to about 55% (w/w)pyrethroid; A) a glycol, glycol ether, glycol ester, fatty acid ester orneutral oil, and B) NMP, wherein said a glycol, glycol ether, glycolester, fatty acid ester or neutral oil, and NMP are present in aweight:weight ratio of about 1:1.8 to about 1:2.8 or about 1:2.0 toabout 1:3.5, about 1:2.0 to about 1:3.0, about 1:2.5 to about 1:3.5 orabout 1:2.5 to about 1:3.0, glycol, glycol ether, glycol ester, fattyacid ester or neutral oil to NMP; and optionally an antioxidant.

A useful concentration of N-arylpyrazole in the composition is fromabout 2% (w/w) to about 15% (w/w). In another embodiment, theconcentration of the N-arylpyrazole is from about 3% (w/w) to about 10%(w/w) or about 4% (w/w) to about 8% (w/w). In still another embodiment,the concentration of the N-arylpyrazole is about 6% (w/w).

A useful concentration of the pyrethroid in the compositions of theinvention is from about 35% (w/w) to about 50% (w/w). In anotherembodiment, the pyrethroid is present at a concentration of from about40% (w/w) to about 48% (w/w). In yet another embodiment, theconcentration of the pyrethroid is from about 42% (w/w) to about 47%(w/w). In another embodiment, the concentration of the pyrethroid in thecomposition is about 45% (w/w).

N-Arylpyrazoles and their parasiticidal and acaricidal activity areknown from US 20060014802 A1, WO2005090313 A1, FR2834288 A1, WO9828277,U.S. Pat. No. 6,069,157, WO0031043, DE19824487, WO9804530, WO9962903,EP0933363, EP0911329, WO9856767, U.S. Pat. No. 5,814,652, WO9845274,WO9840359, WO9828279, WO9828278, DE19650197, WO9824767, EP0846686,EP0839809, WO9728126, EP0780378, GB2308365, U.S. Pat. No. 5,629,335,WO9639389, U.S. Pat. No. 5,556,873, EP0659745, U.S. Pat. No. 5,321,040,EP0511845, and EP0234119, EP0295117, and WO 98/24769. These referencesare hereby incorporated herein in their entirety.

Pyrethroids likewise have a relatively broad parasiticidal action, andsome representatives may also show good acaricidal effects. As discussedabove, the different physicochemical properties of the materials usedrequire special formulations. However, the solvent system describedherein has been shown to solubilize a high concentration of permethrinin combination with fipronil.

In some embodiments the compositions comprising a combination ofN-arylpyrazoles and pyrethroids may contain one or more of acyanopyrethroid (for example flumethrin), a type-1 pyrethroid (forexample permethrin) or non-ester pyrethroid (etofenprox) type. In otherembodiments, the compositions may include one or more of the pyrethroidsα-cyanopyrethroids (for example alpha-cypermethrin, cyfluthrin,beta-cyfluthrin, cyhalothrin, cypermethrin, deltamethrin, fenvalerate,flucythrinate, flumethrin, tau-fluvalinate); type-1 pyrethroids (forexample allethrin, bioallethrin, permethrin, phenothrin, resmethrin,tetramethrin, transfluthrin); and non-ester pyrethroids (for exampleetofenprox, halfenprox, silafluofen).

It is furthermore also possible to use the active compounds in the formof their solvates, in particular hydrates. Solvates are to be understoodas meaning both the solvates, in particular hydrates, of the activecompounds themselves and the solvates, in particular hydrates, of theirsalts.

Also contemplated are the pharmaceutically or veterinarily acceptableacid or base salts, where applicable, of the active compounds providedfor herein. The term “acid” contemplates all pharmaceutically orveterinarily acceptable inorganic or organic acids. Inorganic acidsinclude mineral acids such as hydrohalic acids, such as hydrobromic andhydrochloric acids, sulfuric acids, phosphoric acids and nitric acids.Organic acids include all pharmaceutically or veterinarily acceptablealiphatic, alicyclic and aromatic carboxylic acids, dicarboxylic acids,tricarboxylic acids, and fatty acids. Preferred acids are straight chainor branched, saturated or unsaturated C1-C20 aliphatic carboxylic acids,which are optionally substituted by halogen or by hydroxyl groups, orC6-C12 aromatic carboxylic acids. Examples of such acids are aceticacid, carbonic acid, formic acid, fumaric acid, acetic acid, propionicacid, isopropionic acid, valeric acid, a-hydroxy acids, such as glycolicacid and lactic acid, chloroacetic acid, benzoic acid, methane sulfonicacid, and salicylic acid. Examples of dicarboxylic acids include oxalicacid, malic acid, succinic acid, tartaric acid and maleic acid. Anexample of a tricarboxylic acid is citric acid. Fatty acids include allpharmaceutically or veterinary acceptable saturated or unsaturatedaliphatic or aromatic carboxylic acids having 4 to 24 carbon atoms.Examples include butyric acid, isobutyric acid, sec-butyric acid, lauricacid, palmitic acid, stearic acid, oleic acid, linoleic acid, linolenicacid, and phenylsteric acid. Other acids include gluconic acid,glycoheptonic acid, lactobionic acid, methanesulphonic acid,4-toluenesulphonic acid, galacturonic acid, embonic acid, glutamic acidor aspartic acid.

The term “base” contemplates all pharmaceutically or veterinaryacceptable inorganic or organic bases. Such bases include, for example,the alkali metal and alkaline earth metal salts, such as the lithium,sodium, potassium, magnesium or calcium salts. Organic bases include thecommon hydrocarbyl and heterocyclic amine salts, which include, forexample, the morpholine and piperidine salts.

In one embodiment, the liquid carrier may also comprise a microemulsion.Microemulsions are also well suited as the liquid carrier vehicle.Microemulsions are quaternary systems comprising an aqueous phase, anoily phase, a surfactant and a co-surfactant. They may be translucentand isotropic liquids.

Microemulsions are composed of stable dispersions of microdroplets ofthe aqueous phase in the oily phase or conversely of microdroplets ofthe oily phase in the aqueous phase. The size of these microdroplets isless than 200 nm (1000 to 100,000 nm for emulsions). The interfacialfilm is composed of an alternation of surface-active (SA) andco-surface-active (Co-SA) molecules which, by lowering the interfacialtension, allows the microemulsion to be formed spontaneously.

The oily phase can in particular be formed from mineral or vegetableoils, from unsaturated polyglycosylated glycerides or fromtriglycerides, or alternatively from mixtures of such compounds. Theoily phase may comprise triglycerides including medium-chaintriglycerides, for example C8-C10 caprylic/capric triglyceride. In someembodiments the oily phase will represent, in particular, in aconcentration of about 2 to about 15%, about 7 to about 10%, or fromabout 8 to about 9%, v/v of the microemulsion.

In some embodiments, the aqueous phase includes, for example water orglycol derivatives, such as propylene glycol, glycol ethers,polyethylene glycols or glycerol. Propylene glycol, diethylene glycolmonoethyl ether and dipropylene glycol monoethyl ether are especiallypreferred. Generally, the aqueous phase will represent a proportion fromabout 1 to about 4% v/v in the microemulsion.

Surfactants for the microemulsion include diethylene glycol monoethylether, dipropyelene glycol monomethyl ether, polyglycolized C8-C10glycerides or polyglyceryl-6 dioleate. In addition to these surfactants,the co-surfactants include short-chain alcohols, such as ethanol andpropanol.

Some compounds are common to the three components discussed above, e.g.,aqueous phase, surfactant and co-surfactant. However, it is well withinthe skill level of the practitioner to use different compounds for eachcomponent of the same formulation.

In one embodiment, the co-surfactant to surfactant ratio will be fromabout 1/7 to about 1/2. In another embodiment, the concentration ofsurfactant will be from about 25 to about 75% v/v and the concentrationof the co-surfactant will be from about 10 to about 55% v/v in themicroemulsion.

Likewise, the co-solvents are also well known to a practitioner in theformulation art. In some embodiments, co-solvents are those that promotedrying and include, for example, absolute ethanol, isopropanol(2-propanol) or methanol.

For the chemical preparation of the products of the invention, a personskilled in the art is regarded as having at his or her disposal, interalia, the entire contents of “Chemical Abstracts” and of the documentsthat are cited therein.

Depending on the nature and arrangement of the substituents, the activecompounds may, if appropriate, be present in various stereoisomericforms, in particular as enantiomers and racemates. According to theinvention, it is possible to use both the pure stereoisomers andmixtures thereof.

The present subject matter is further described herein by the followingnon-limiting examples which further illustrate the invention, and arenot intended, nor should they be interpreted to, limit the scope of theinvention.

EXAMPLES Example 1: Stable Fly Repellency and Parasiticidal Efficacy inRats

This study evaluated the effectiveness of a formulation of the inventioncontaining fipronil and permethrin to repel and kill Stomoxys calcitranson rats after a single administration. Rats in the treatment group(Group 2) were treated once on Day 0 with a topical formulation of theinvention containing 9.2 (w/w) fipronil and 41.5% (w/w) permethrin in asolvent system containing DMDA and DGME in a ratio of 0.73:1, DMDA toDGME, to deliver a dose of 30 mg/kg permethrin and 6.7 mg/kg fipronil.Rats in Group 3 were treated with a formulation containing permethrinalone to deliver a dose of 30 mg/kg. The two treatment groups werecompared with a control group treated with a solvent system placebo.

Flies are exposed to the treated rats for 1 hr., then removed. Deadflies are counted at the end of the exposure period and at 4 hr and 24hr post-exposure. The fipronil+permethrin group exhibited a very highlevel of efficacy at all time points measured. Furthermore, thefipronil+permethrin group exhibited significantly higher repellencymeasured 24 hours after exposure compared with the permethrin only groupat the same dose level. This finding is significant because fipronil isnot known to have repellent activity.

The diptericidal efficacy of the formulations is shown in Table 1 andrepresented in FIG. 6. The data for repellency are shown in Table 2 andare represented in FIG. 7.

TABLE 1 Dose EOE 4 hr 24 hr Treatment (mg/kg) % efficacy % efficacy %efficacy Permethrin 30.0 74.2 82.0 74.7 Fipronil + 6.7 95.8 95.7 95.6Permethrin 30.0 EOE = end of exposure period

TABLE 2 Treatment Dose (mg/kg) % Repellency Permethrin 30.0 41.2Fipronil + 6.7 55.9 Permethrin 30.0

Example 2: Stable Fly Repellency and Parasiticidal Efficacy on Dogs

This study evaluated the effectiveness of Fipronil+permethrin to repeland kill Stomoxys calcitrans on dogs after a single administration.

The fly repellency and parasiticidal activity of a formulation of theinvention was evaluated on ten dogs. Dogs were⁻ randomly allocated toone of two groups by lottery. Dogs in Group 1 were not treated: Dogs inGroup 2 received the topical formulation of the invention containing acombination of fipronil and permethrin described in Table 3 below,administered topically once on Day 0 in a single spot to deliver a doseof approximately 6.70 mg fipronil/kg and 30.15 mg permethrin/kg.

TABLE 3 Formulation Ingredient % w/v % w/w fipronil 10 9.2 Permethrin 4541.5 Dimethyl decanamide QS ~49 (DMDA):Diethylene glycol monoethyl ether(DGME) at 0.7:3:1 (w/w)

On Days 1, 14, 21 and 28, dogs were exposed to approximately 100 threeto seven day old S. calcitrans. After approximately 60 minutes, liveflies were aspirated into a vial and dead flies were collected from thecages. After all flies were collected, the live flies were killed andthe flies (live and dead) were counted and crushed to determine feedingstatus (fed vs. unfed). Due to inadequate fly emergence, fly exposurescould not be conducted on Day 7

Fly repellency was based on the number of flies that fed in the treatedgroup compared to the control group during each exposure period.Repellency was 99% in treated dogs on Day 1 and was 85.9%, 84.6%, and61.7% on Days 14, 21, and 28, respectively.

Parasiticidal efficacy was based on the number of live flies in thetreated group compared to the control group at the end of each exposureperiod. Parasiticidal efficacy in treated dogs was 100% on Day 1 and was98.7%, 95.5%, and 81.3% on Days 14, 21 and 28, respectively. The percentefficacy is listed in Table 3.

In this study, a composition comprising Fipronil+permethrin provided≥84.6°/0 repellency and ≥95.5% parasiticidal efficacy against S.calcitrans (stable flies) for three weeks following treatment.

Summary Data are shown for the fipronil+permethrin composition in Table4:

TABLE 4 Summary data Day 1 7 14 21 28 Repellency 99.0 N/A 85.9 84.6 61.7Parasiticidal 100.0 N/A 98.7 95.5 81.3 Activity Example 3:Crystallization of Actives

Four experimental formulations were prepared with 6% w/w Fipronil and44.9% w/w permethrin and varying amounts of excipientsN-dimethyldecanamide (DMDA) and diethylene glycol monoethyl ether(DGME). The DMDA concentration was varied between 5-20% w/w. Theseformulations were subjected to different low temperature conditionsincluding −20° C., 4° C., and 10° C.; some samples were seeded withcrystals that had grown in previous batches. Crystallization was seenamong all the formulations in various conditions.

A DMDA-containing formulation of fipronil and permethrin as describedherein was subjected to low temperatures. Some samples were seeded withcrystals. Crystals were seen in the DMDA formulation.

Solvents were screened for their compatibility with permethrin andability to maintain permethrin in solution. The summary of the testingand results are shown in Table 5.

TABLE 5 Solvent Crystals Captex 170 (Caprylic/Capric acid 100% Ester)Oleic acid 100% Propylene glycol Immiscible Benzyl benzoate Less than 5%Propylene carbonate 100% Glycerol formal 100% Ethanol  40% Isopropylalcohol Immiscible Diethylene glycol monoethyl ether (DGME) 100%N,N-Dimethyldecanamide (DMDA) Less than 5%

The screening indicates benzyl benzoate and N,N-Dimethyldecanamide aremost compatible with permethrin from the experimental set.

In addition to screening solvents for their compatibility withpermethrin, several experimental batches were prepared all containing44.9% w/w permethrin and 6% w/w fipronil. The results of thepreparations are shown in Tables 6 and 7.

TABLE 6 Solvent Result DGME, ethanol, seeded and unseeded samples storedat 4° C. polysorbate 80, PVP crystallized 100% within 24 hoursN-methylpyrrolidone seeded and unseeded samples stored at 4° C. have notgrown crystals after 5 days Dimethyl Isosorbide/ Solution turned hazyafter all ingredients Diisopropyl adipate added DimethylIsosorbide/Benzyl Fipronil was insoluble benzoate Triacetin/diisopropylFipronil was insoluble adipate Triacetin/benzyl benzoate Fipronil wasinsoluble

The formulation containing N-methylpyrrolidone has the best physicalstability of the set.

TABLE 7 Formulation (% w/w or grams per 100 g of solution)Crystallization on day 8 Prepared Without Solvent system FipronilPermethrin on seeding With seeding DGME 24.5 6 45 Day 1 No YesDiisopropyl Adipate 24.5 Polysorbate 80 3.0 6 45 Day 1 No Yes DGME 23.0Diisopropyl Adipate 23.0 DGME 32 6 45 Day 2 No Yes Diisopropyl Adipate17 DMDA 23.5 6 45 Day 3 No Yes Diisopropyl Adipate 25.5 DMDA 16 6 45 Day3 No Yes Diisopropyl Adipate 33 NMP 14.5 6 45 Day 4 No Yes, very smallDiisopropyl Adipate 34.5 amounts of crystals NMP 49 6 45 Day 5 No No NMP44 6 45 Day 5 No No Diisopropyl Adipate 5 NMP 44 6 45 Day 5 No NoTriacetin 5 Note: 1) These results are approximate and based on labscreening data. 2) Amount of DGME, DMDA, NMP is approximate and willdepend on assay of actives. 3) Assay of all actives is assumed 100% inthese experiments and calculations.

Example 4: One Formulation of the Invention Described Fully Herein isShown in Table 8

TABLE 8 Ingredient % w/w % w/v NMP 35 39.4 BHT 0.10 0.11 Fipronil 6.06.8 Permethrin 44.9 50.5 MIGLYOL ® 812 QS (~14) QS (~15)

Example 5: Efficacy of Against Fleas (Ctenocephalides felis) and Ticks(Rhipicephalus sanguineus) on Dogs

A study was conducted to evaluate the effectiveness of three differentformulations of the invention containing fipronil and permethrin inamounts to deliver doses of ≥6.7 mg/kg and ≥50.3 mg/kg, respectively,compared with a formulation containing permethrin alone in an amount todeliver a dose of ≥50.3 mg/kg.

Five treatment groups of six dogs each were formed. Dogs in Group 1 wereuntreated. On Day 0 dogs in Groups 2, 3 and 4 received a spot-onformulation of the invention containing 6.0% (w/w) fipronil and 44.9%(w/w) permethrin in different solvent systems. Dogs in Group 5 weretreated on Day 0 with a spot-on formulation containing 44.9% (w/w)permethrin alone. The efficacy of the treatments was measured againstthe control Group 1 for each observation point. The formulationcomponents are shown in Table 9 below.

TABLE 9 Group 2 Group 3 Group 4 Group 5 Ingredient (% w/w) (% w/w) (%w/w) (% w/w) Fipronil 6.0 6.0 6.0 Permethrin 44.9 44.9 44.9 44.9 NMP 3535 35 DGME QS (~12.5) QS (~18.7) MIGLYOL ® QS (~12.4) 812 BCA 25 DMDA QS(~22.6) BHT 0.1 0.1 0.1

All dogs were infested with approximately 100 C. felis on Days −1, 8,15, 22 and 29. Flea counts were performed on Days 1, 9, 16, 23 and 30.All dogs were also infested with approximately 50 Rhipicephalussanguineus on Days −1, 14 and 28. Ticks were thumb-counted on Days 15and 29 and counted and removed on Days 1, 16 and 30.

The efficacy against fleas was similar for Groups 2, 3 and 4, rangingfrom 95-100% through Day 30. The efficacy for Group 5 was 51% on Day 1and reached a peak of 96% on Day 9, then dropped to 13% by Day 30.

The efficacy against ticks at 24 hours after treatment was 69.1%, 89.9%,94.7% and 70.2% for Groups 2, 3, 4 and 5, respectively. Efficacy againstticks at 24 hours following infestation ranged from 91-99% across alltreated groups on Day 15 and from 56-81% on Day 29. Efficacy againstticks at 48 hours after infestation ranged from 96-100% across alltreated groups on Day 16 and from 74-85% on Day 30.

This study demonstrates that the formulations of the invention providedsuperior efficacy against C. felts and R. sanguineus compared to aformulation containing permethrin alone. The efficacy data is alsopresented in FIGS. 4 and 5.

Example 6: Repellency and Efficacy Against Phlebotomus perniciosus SandFlies on Dogs

This study evaluated the effectiveness of two formulations of theinvention containing fipronil and permethrin to repel and killPhlebotomus perniciosus on dogs after a single administration.

Following a procedure very similar to that described in Example 2, therepellency and insecticidal efficacy of the formulations was studied.Three treatment groups of five dogs each were formed. Dogs in Group 1were untreated. Dogs in Groups 2 and 3 were treated once on Day 0 withthe formulations of the invention described in Table 10 below to deliverdoses of ≥6.7 mg/kg fipronil and ≥50.3 mg/kg permethrin.

TABLE 10 Group 2 Group 3 Ingredient (% w/w) (% w/w) Fipronil 6.0 6.0Permethrin 44.9 44.9 NMP 35 35 DGME QS (~12.5) MIGLYO ®L 812 QS (~12.4)BCA DMDA BHT 0.1 0.1

Dogs were exposed to eighty (±5) P. perniciosus female sand flies onDays 1, 7, 14, 21, 29 and 35. After 60 minutes, sand flies were removedand categorized as live (engorged or non-engorged), dead (engorged ornon-engorged). Efficacy was determined by determining the number of deadflies approximately four hours after exposure and on days 2, 8, 15, 22and 29 approximately twenty four hours after exposure. Repellency wasmeasured by comparing the number of engorged flies (live or dead) in thetreatment groups compared with the number of engorged flies in thecontrol group.

The repellency observed for dogs in treatment Group 2 on days 1, 7, 14,21, 29 and 35 was measured to be 97.0%, 90.1%, 96.0%, 80.0%, 96.4% and79.6%, respectively. The repellency for dogs in Group 3 on days 1, 7,14, 21, 29 and 35 was found to be 94.4%, 94.7%, 99.3%, 88.6%, 97.3% and82.5%, respectively.

The efficacy for Group 2 at 4 hours after exposure on days 1, 7, 14, 21,29 and 35 was 99.8%, 100.0%, 94.0%, 63.2%, 91.1% and 65.8%,respectively. The efficacy for Group 3 at 4 hours after exposure on days1, 7, 14, 21, 29 and 35 was 99.6%, 100.0%, 92.3%, 56.3%, 90.6% and70.0%, respectively.

The efficacy for Group 2 at 24 hours after exposure on days 2, 8, 15,22, 30 and 36 was 100.0%, 99.4%, 94.4%, 66.3%, 91.7% and 69.6%,respectively. The efficacy for Group 3 at 4 hours after exposure on days2, 8, 15, 22, 30 and 36 was 99.6%, 100.0%, 92.6%, 58.0%, 93.1% and70.7%, respectively.

This study demonstrates that the formulations of the invention provideexcellent repellency for at least 35 days and excellent insecticidalefficacy for at least 30 days against Phlebotomus perniciosus on dogsafter a single treatment. The data is also presented in FIG. 3.

Example 7: Repellency and Efficacy Against Stable Flies (Stomoxyscalcitrans) on Dogs

This study was conducted to evaluate the repellency and efficacy of aspot-on formulation of the invention described in Table 8 abovecomprising a combination of fipronil and permethrin against Stomoxyscalcitrans stable flies after a single topical administration.

Following a procedure very similar to that described in Example 2 above,two groups of eight dogs each were formed. The dogs in Group 1 wereuntreated and served as a control group. Dogs in Group 2 were treatedonce on Day 0 with the formulation of Table 8 to deliver doses of 6.8mg/kg fipronil and 50.5 mg/kg permethrin. Dogs were exposed toapproximately 100 stable flies on days 1, 7, 14, 21, 28 and 35 forapproximately 60 minutes, after which the flies (both live and dead)were collected and examined to determine feeding status. Repellency ofthe formulation administered to Group 2 dogs was measured to be 100.0%,100.0%, 99.2%, 97.3%, 96.6% and 88.7% on days 1, 7, 14, 21, 28 and 35,respectively. The insecticidal efficacy of the formulation applied toGroup 2 dogs was measured to be 100.0%, 99.9%, 99.4%, 99.6%, 98.3% and98.3% on Days 1, 7, 14, 21, 28 and 35 respectively. The repellency andefficacy data is presented in FIGS. 8 and 9.

The very high level of repellency and insecticidal efficacy againstStomoxys calcitrans with a duration of action of at least 35 days forthe formulation of Example 8 is surprising for a topical formulationcomprising permethrin at the dose level applied. For example, Fourie etal. reported that a topical formulation comprising 10% imidacloprid and50% permethrin (the same concentration of permethrin in the formulationused in the present example) only prevented 82% of S. calcitrans fromfeeding on dogs for 4 weeks after treatment and exhibited an averageefficacy of 85.6% over the evaluation period of 29 days. (see “TheEfficacy of a Topically Applied Combination of Imidacloprid andPermethrin Against Stomoxys calcitrans on Dogs,” Intern. J. Appl. Res.Vet. Med., 2006, vol. 4(1), pp. 29-33). In this study, the highestrepellency observed over the whole 29 day assessment period was 90.2% 1day post treatment while the repellency at all other time points werebelow 90%. The repellency and efficacy exhibited by the formulation ofthe invention is surprisingly superior to another topical formulationcomprising the same concentration of permethrin in combination with adifferent pesticidal active agent. The superior repellency isparticularly surprising because fipronil is not known to have anyrepellent activity.

Example 8: Repellency Against Mosquitoes (Culex pipiens) on Dogs

This study was conducted to evaluate the repellency and efficacy of aspot-on formulation of the invention described in Table 8 above againstCulex pipiens after a single topical administration.

Following a procedure very similar to that described in Example 2 above,two groups of eight dogs each were formed. The dogs in Group 1 wereuntreated and served as a control group. Dogs in Group 2 were treatedonce on Day 0 with the formulation of Table 8 to deliver doses of 6.8mg/kg fipronil and 50.5 mg/kg permethrin. Dogs were exposed toapproximately 100 Culex pipiens female mosquitoes per dog on days 1, 7,14, 21 and 28 for approximately 60 minutes, after which the mosquitoes(both live and dead) were collected and examined to determine feedingstatus.

Repellency of the formulation administered to Group 2 dogs was measuredto be 99.4%, 98.9%, 94.7%, 91.7% and 90.4% for days 1, 7, 14, 21 and 28,respectively. The repellency data is presented in FIG. 10.

Throughout this specification and the claims, the words “comprise,”“comprises,” and “comprising” are used in a non-exclusive sense, exceptwhere the context requires otherwise.

As used herein, the term “about,” when referring to a value is meant toencompass variations of, in some embodiments ±20%, in some embodiments±10%, in some embodiments ±5%, in some embodiments ±1%, in someembodiments ±0.5%, and in some embodiments ±0.1% from the specifiedamount, as such variations are appropriate to perform the disclosedmethods or employ the disclosed compositions.

All publications, patent applications, patents, and other references areherein incorporated by reference to the same extent as if eachindividual publication, patent application, patent, and other referencewas specifically and individually indicated to be incorporated byreference. It will be understood that, although a number of patentapplications, patents, and other references are referred to herein, suchreference does not constitute an admission that any of these documentsforms part of the common general knowledge in the art.

Although the foregoing subject matter has been described in some detailby way of illustration and example for purposes of clarity ofunderstanding, it will be understood by those skilled in the art thatcertain changes and modifications can be practiced within the scope ofthe appended claims.

Having thus described in detail certain embodiments of the presentinvention, it is to be understood that the invention defined by theabove paragraphs is not to be limited to particular details set forth inthe above description as many apparent variations thereof are possiblewithout departing from the spirit or scope of the present invention.

That which is claimed:
 1. A topical composition for repelling anectoparasite on an animal comprising: from about 2% (w/w) to about 15%(w/w) fipronil; from about 30% (w/w) to about 55% (w/w) permethrin; anda neutral oil(s) and N-methyl pyrrolidone, wherein said neutral oil(s)and N-methyl pyrrolidone are present in a weight:weight ratio of fromabout 1:2.0 to about 1:3.5, neutral oil(s) to N-methyl pyrrolidone andwherein the amounts of fipronil and permethrin in the neutral oil andN-methylpyrrolidone are in repellently effective amounts to repel theectoparasite when the composition is applied to the skin of the animal.2. The composition of claim 1, wherein said fipronil is present in aconcentration of from about 4% (w/w) to about 8% (w/w).
 3. Thecomposition of claim 1, wherein said fipronil is present at aconcentration of about 6% (w/w).
 4. The composition of claim 1, whereinsaid permethrin is present in a concentration of from about 35% (w/w) toabout 50% (w/w).
 5. The composition of claim 1, wherein said permethrinis present in a concentration of from about 40% (w/w) to about 48%(w/w).
 6. The composition of claim 1, wherein said permethrin is presentat a concentration of about 45% (w/w).
 7. The composition of claim 1,wherein said fipronil is present at a concentration of about 6.0% (w/w)and said permethrin is present at a concentration of about 45% (w/w). 8.The composition of claim 1, wherein said neutral oil(s) and saidN-methyl pyrrolidone are present in a weight:weight ratio of about 1:2.0to about 1:3.0, neutral oil(s) to N-methyl pyrrolidone.
 9. Thecomposition of claim 1, wherein said neutral oil(s) is present in aconcentration of about 12% (w/w) to about 14% (w/w) and said N-methylpyrrolidone is present in a concentration of about 35% (w/w).
 10. Thecomposition of claim 1, wherein said neutral oil(s) and said N-methylpyrrolidone are present in a weight:weight ratio of about 1:2.5 to about1:3.0, neutral oil(s) to N-methyl pyrrolidone.
 11. The composition ofclaim 1, wherein said neutral oil(s) and said N-methyl pyrrolidone arepresent in a weight:weight ratio of about 1:2.5 to about 1:3.5, neutraloil(s) to N-methyl pyrrolidone.
 12. The composition of claim 1, whereinsaid neutral oil(s) and said N-methyl pyrrolidone are present in aweight:weight ratio of about 1:2.2 to about 1:2.4, neutral oil(s) toN-methyl pyrrolidone.
 13. The composition of claim 1, wherein saidneutral oil(s) and said N-methyl pyrrolidone are present in aweight:weight ratio of about 1:2.5 to about 1:2.8, neutral oil(s) toN-methyl pyrrolidone.
 14. The composition of claim 1, wherein saidneutral oil(s) is a triglyceride of fractionated plant fatty acidshaving chain lengths of C₈ and C₁₀.
 15. The composition of claim 1,comprising about 6% (w/w) fipronil; about 45% (w/w) permethrin; about12% to about 14% (w/w) of triglycerides of fractionated plant fattyacids having chain lengths of C₈ and C₁₀ (w/w); about 35% (w/w) N-methylpyrrolidone; and about 0.1% (w/w) butylated hydroxytoluene.
 16. Thecomposition of claim 1, further comprising an antioxidant.
 17. Thecomposition of claim 16, wherein said antioxidant is butylatedhydroxytoluene.
 18. The composition of claim 1 further comprising one ormore additional active agents.
 19. The composition of claim 18, whereinthe one or more additional active agent is an avermectin, a milbemycin,a spinosyn, a spinosoid, a benzimidazole, levamisole, pyrantel,morantel, praziquantel, closantel, clorsulon, an amino acetonitrileactive agent, an insect growth regulator, a neonicotinoids or anaryloazol-2-yl cyanoethylamino active agent, or a combination ofthereof.
 20. A spot-on composition for repelling an ectoparasite on ananimal, comprising from about 2% (w/w) to about 15% (w/w) fipronil; fromabout 30% (w/w) to about 55% (w/w) permethrin; and a neutral oil(s) andN-methyl pyrrolidone, wherein said neutral oil(s) and N-methylpyrrolidone are present in a weight:weight ratio of from about 1:2.5 toabout 1:3.5, wherein said composition is a liquid having a volume offrom about 1 mL to about 10 mL; and wherein the amounts of fipronil andpermethrin in the neutral oil and N-methylpyrrolidone are in repellentlyeffective amounts to repel the ectoparasite when the composition isapplied to the skin of the animal.
 21. The composition of claim 1 or 20,further comprising fipronil sulfone below about 3.5% by area relative tothe peak area for fipronil as measured by HPLC at about three monthsafter formulation.
 22. The composition of claim 1 or 20, furthercomprising fipronil sulfone wherein said amount of fipronil sulfone atabout three months after formulation has not increased by more than 50%of the original amount of said fipronil sulfone present at the time offormulation.
 23. (canceled)
 24. A method for repelling an ectoparasiteon an animal, comprising administering an effective amount of thecomposition of claim 1 to the animal.
 25. The method of claim 24,wherein said administering comprises contacting the coat and/or skin ofsaid animal with said composition.